Post on 29-Jan-2023
Abstracts 3rd Biennial Meeting of InSiGHT(International Society for Gastrointestinal Hereditary Tumours)
Dusseldorf June 24–27 2009
Small bowel polypectomy by double balloon
enteroscopy: advancing endoscopic therapy for patients
with Peutz–Jegher’s syndrome
Edward Despott, Robin Phillips, Sue Clark, Aymer Postgate, Aine
Fitzpatrick, Eric Tripoli, Krysia Konieczko, Chris Fraser
The Wolfson Unit for Endoscopy & The Polyposis Registry,
St Mark’s Hospital, Harrow, UK
Introduction Small bowel (SB) obstruction and GI bleeding due to
polyps are major causes of morbidity in patients with Peutz–Jegher’s
Syndrome (PJS). Priorto double balloon enteroscopy (DBE), a tech-
nique that allows complete enteroscopy, the main therapeutic option
available for PJS patients was laparotomy with intra-operative enter-
oscopy. We report our experience of the endoscopic management by
DBE of SB polyps in the largest cohort of PJS patients from the UK.
Aims & methods Data on PJS cases managed by DBE at St Mark’s
since 2005 were prospectively collected. Patients deemed to have
significant SB polyps ([15 mm) at capsule endoscopy or magnetic
resonance enterography or both, were referred for elective DBE
polypectomy. DBE procedures were performed under GA. In one
case, an emergency DBE was performed in the setting of SB
obstruction due to polyp-induced intussusception. All patients remain
under follow up and surveillance.
Results Nine patients (mean age 34 years, range 16–45 years)
underwent 13 DBE procedures (3 patients had 2 DBEs). Although all
patients had previous laparotomies (some multiple), adhesions did not
cause significant hindrance to DBE. On average 3 polyps were
removed per patient (mean size 18 mm, range 8–37 mm). Polyp
stalks were injected with lifting solution (dilute adrenaline and
methylene blue) with additional endo-looping on occasion priorto
snaring to reduce bleeding risk. Sessile polyps were elevated with
lifting solution to minimise perforation risk. One patient required 3
DBE procedures, one of which was laparoscopically assisted fora
sessile distal duodenal polyp; 1 patient suffered a small post-poly-
pectomy bleed which settled spontaneously and 1 patient in whom an
emergency DBE was attempted, polypectomy of a sessile 30 mm
polyp led to perforation with conversion to IOE through the defect.
Conclusion We highlight the role of DBE as an emerging thera-
peutic option for SB polyps in PJS that avoids the need for
laparotomy and prolonged recovery, however experienced surgical
cover should always be available for this complex group of patients in
view of the risk of potential complications.
High cumulative risk of intussusceptions in patients
with Peutz–Jeghers syndrome
M. G. F. van Lier, A. Wagner, A. M. Westerman, J. H. P. Wilson,
F. W. M. de Rooij, E. J. Kuipers, M. E. van Leerdam
Erasmus MC, University Medical Center, Rotterdam,
The Netherlands
Peutz–Jeghers Syndrome (PJS) is an inherited disorder characterized
by gastrointestinal hamartomas and mucocutaneous pigmentations.
Germline mutations in the STK11-gene can be found in 70% of
clinically affected patients. Hamartomas, mainly located in the small
bowel, may cause intussusceptions. Since balloon-enteroscopy (BE)
enables endoscopic removal of these polyps, we assessed the risk and
onset of intussusception.
Patients diagnosed with PJS based on clinical diagnostic criteria or
proven STK11 mutation were included in this prospective cohort study
(1995–2008). Clinical data were obtained by interview and chart-
review. Genotype-phenotype correlations were evaluated. The cumu-
lative risk of intussusception was calculated by Kaplan–Meier analyses.
Forty-four PJS patients (57% males) were included from 18 PJS
families; 32 patients still alive had a median age of 44 years
(10–74 years) and 12 patients had deceased at a median age of
45 years (11–73 years). A germline STK11 mutation was detected in
32 patients (73%). Thirty-four patients (77%) had a history of one or
more (1–6) episodes of intussusception due to small bowel polyps.
The median age at the first intussusception was 13.5 years
(3–50 years). Surgery was required in 33 patients (75%). There was
no significant difference in intussusception incidence according to sex
(p = 0.15) or mutation-status (p = 0.70). Kaplan–Meier analyses
showed that intussusception had occurred in 50% of the cohort at a
median age of 16 years (95% CI 11–21), increasing to 75% (95% CI
62–88) at the age of 35 years. The 10-year probability of intussus-
ception was 25% (95% CI 12–38).
PJS patients carry a high cumulative risk of intussusception caused
by small bowel hamartomas (50% at 16 years), independent of
STK11 mutation-status. These findings support the approach of
123
Familial Cancer (2010) 9:713–748
DOI 10.1007/s10689-010-9351-8
enteroscopic surveillance with removal of small bowel hamartomas.
The effect on the incidence of intussusception remains to be estab-
lished and weighted against burden and complication-risk of the
intervention.
Phenotype variability within CDH1 mutated families
Laetitia Huiart1, Francois Eisinger1, Violaine Bourdon1, Bruno
Buecher2, Martine Blayau2, Olivier Caron2, Jean-Francois Flejou2,
Jean-Pierre Gendre2, Astrid Schielke2, Alain Sezeur2
1Sylviane Olschwang pour le reseau PHRATries;2Institut Paoli-Calmettes, Marseille, France
CDH1 mutations are associated with diffuse gastric carcinoma and
lobular breast cancer. However, little is known on age dependent
penetrance of CDH1 mutations and on relevance of current clinical
management guidelines. Our objective was therefore to describe
personal and familial history of cancer in families with a CDH1
mutation.
We identified 16 families who tested positive for a deleterious
CDH1 mutation between 1998 and 2008. All but one mutation were
unique: c.1565+1del was found in 4 unrelated families. Index cases
were affected with gastric cancer in all cases but 2. Nine out of the
14 index cases of gastric cancers were diffuse, 5 were unspecified.
Ages at diagnosis ranged from 21 to 63 years (mean = 38). The 2
index cases free of gastric cancers were tested either because of a
bilateral lobular breast cancer at 42, or because of a rectal linitis at
23 years of age. Overall 35 cases of gastric cancers were reported.
No other family history of gastric cancer was found for 5 mutation
carriers. Associated breast cancer was reported in 3 families, of
which 2 were specified as lobular breast cancer. Interestingly in a
family where 13 members were tested for the familial mutation
(4 carriers, 9 non carriers), 2 mutation carriers were free of cancer at
65 and 49 respectively. In another family, among 10 mutation
carriers, 6 underwent prophylactic gastrectomy, all showed invasive
and in situ signet cell foci. However the oldest carrier who declined
gastrectomy was clinically asymptomatic at 64 years of age. In total,
6 mutation carriers were clinically asymptomatic although older
than the mean age at diagnosis observed in index cases. The int-
rafamilial phenotype variability observed in our series indicates the
need for international consortium to provide reliable data on pene-
trance of CDH1 mutations before validate guidelines for clinical
management.
Risk of pancreatic cancer in hereditary nonpolyposis
colorectal cancer
Jennifer E. Axilbund1, Alison P. Klein1,2,3, Judith A. Bacon2,
Li Wang3, Daniel Edelstein4, Carolina E. Fasola2, Francis M.
Giardiello1,4, Constance A. Griffin1,2
1Department of Oncology, 2Department of Pathology, Sol Goldman
Pancreatic Cancer Research Center, 4Department of Medicine, The
Johns Hopkins University School of Medicine, Baltimore, MD, USA
Background Hereditary Nonpolyposis Colorectal Cancer (HNPCC)
is characterized by early-onset colorectal cancer. Other associated
cancers include endometrial, ovarian, gastric, small bowel, urinary
tract and biliary tract. It has been suggested that the risk of pancreatic
cancer is also increased, though the precise magnitude is unknown.
Pancreatic cancer is the 4th leading cause of cancer death in the
United States, leading to an estimated 35,000 annual deaths. The aim
of this study was to assess the lifetime risk of pancreatic cancer in
HNPCC families to guide screening recommendations.
Methods In a retrospective, registry and clinic questionnaire-based
study, we estimated the risk of pancreatic cancer in families that
meet the Amsterdam-I criteria and/or have an identified mutation in
an HNPCC-associated gene. Families were specifically queried
regarding pancreatic cancer occurrence to avoid bias in reporting to
a colon cancer registry and/or clinic. Person-years of follow-up
were calculated for each individual from their birth until either the
date health information was last updated, date of pancreatic cancer
diagnosis or death. Standardized incidence ratios (SIR) were cal-
culated by comparing the observed number of pancreatic cancer
cases among individuals enrolled in the Johns Hopkins Hereditary
Colorectal Cancer Registry and/or the Johns Hopkins Cancer
Risk Assessment Program to those expected from SEER incidence
rates.
Results 1,045 individuals from 66 families were followed for a total
of 63,045 person-years. 8 pancreatic cancers were reported. Overall
risk of pancreatic cancer was 1.38 (CI = 0.63–2.63.) Risk of pan-
creatic cancer in individuals with a first-degree relative with
colorectal or endometrial cancer was 1.51 (CI = 0.65–2.97).
Conclusions These RESULTS: (a) demonstrate a small, not statis-
tically significant increased risk for pancreatic cancer with HNPCC,
and (b) suggest that pancreatic cancer screening is not currently
warranted for the majority of HNPCC families.
Funded by the Jennifer L. Brager Memorial Fund and CA62924.
Results of surveillance for hereditary pancreatic cancer
using annual MRI (CP)
Wouter H. de Vos1, Martin N. Wasser2, Bert A. Bonsing3, Anneke M.
van Mil4, Daniel W. Hommes1, Johan A. Offerhaus6, Hans Morreau5,
Hans F. Vasen1,7
Department of 1Gastroenterology and Hepatology, 2Radiology,3Surgery, 4Clinical Genetics, 5Pathology, Leiden University Medical
Center, Leiden, The Netherlands; 6Pathology Utrecht University
Medical Center, Utrecht, The Netherlands; 7Netherlands Foundation
for the Detection of Hereditary Tumors, Leiden, The Netherlands
Surveillance for pancreatic cancer (PC) in high risk groups may lead
to early detection and may improve overall survival. We screened for
early pancreatic neoplasia in individuals with a p16-germline muta-
tion or with a strong family history of PC (FPC).
Methods Since 2000, high risk individuals were offered annual
surveillance by Magnetic Resonance Imaging (MRI(CP)). In case of
suspected lesions surgery or additional follow up (FU) was pro-
vided. In case of doubt, the examination was repeated within
2–4 months.
Results 71 (28 males) individuals with an average age of 56 years
(range 40–72) were studied (64 from p16-and 7 from FPC families).
The median FU was 3.4 years (range 0–7). IPMN (intraductal pap-
illary mucinous neoplasm)-like lesions were identified in at least five
p16 carriers (8%) and in four patients from FPC families (57%).
Three patients (FPC n = 2 and p16 n = 1) underwent a prophylactic
partial resection of the pancreas. Six patients (9%) were diagnosed
with PC. Of them, 4 asymptomatic patients underwent a partial
pancreatectomy. Two of 4 had no evidence of residual disease after
surgery. The third patient had positive resection margins and nodes
but survived 2 years. The fourth patient had both metastatic carcinoid
714 Abstracts
123
and pancreatic cancer at surgery. The remaining two patients did not
undergo surgery because of metastatic disease; one had pulmonary
metastases of a melanoma, the other patient developed metastatic
pancreatic cancer 3 months after the first screening examination on
which a small pancreatic tumor was missed.
Conclusion Small premalignant lesions can be identified by
MRI(CP). In view of the substantial morbidity and mortality of
pancreatic surgery, a major challenge is to decide at what stage and to
what extent a patient should undergo prophylactic surgery. Close
observation of patients with pancreatic lesions could add valuable
information to this question.
Comparative yield of endosonography and magnetic
resonance imaging in individuals at high-risk
for pancreatic cancer
F. Harinck1, I. Kluijt6, J.-W. Poley1, A. Cats7, C. M. Aalfs4,
D. J. Gouma3, C. Y. Nio5, P. Fockens2, M. J. Bruno1,2
1Department of Gastroenterology and Hepatology, Erasmus Medical
Center Rotterdam, Netherlands; 2Department of Gastroenterology and
Hepatology, Academic Medical Center Amsterdam, Netherlands;3Department of Surgery, Academic Medical Center Amsterdam,
Netherlands; 4Department of Clinical Genetics, Academic Medical
Center Amsterdam, Netherlands; 5Department of Radiology,
Academic Medical Center, Amsterdam Netherlands; 6Department
of Clinical Genetics, Netherlands Cancer Institute Amsterdam,
Netherlands; 7Department of Gastroenterology and Hepatology,
Netherlands Cancer Institute Amsterdam, Netherlands
Introduction Individuals at high-risk for pancreatic cancer (PC) are
(1) mutation carriers of PC prone hereditary syndromes and (2) first-
degree relatives of patients with PC from familial PC kindreds. Non-
invasive pre-cursor lesions of PC include pancreatic intraepithelial
neoplasia (PanIN) and intraductal papillary mucinous neoplasia
(IPMN). A surveillance program may improve the prognosis of high-
risk individuals by detecting asymptomatic early cancers or precursor
lesions. Endosonography (EUS) has shown to be a potentially valu-
able tool. Data for MRI are lacking. We present preliminary results of
a comparative study between baseline EUS and MRI screening
investigations in individuals entering a yearly surveillance program.
Methods Asymptomatic high-risk individuals prospectively under-
went EUS and MRI. Both investigations were carried out and scored
according to predefined criteria. Investigators were blinded to the
results of the alternative imaging modality.
Results Thirty-three individuals underwent both EUS and MRI. In
eight individuals (24%) focal lesions were detected, one with a mass
lesion (11 mm) and seven with cystic lesions. The mass lesion, latter
proven to be an adenocarcinoma, was only detected by EUS. The
cystic lesions were detected by both techniques in four individuals
(12%), by MRI only in two (6%) and by EUS only in two (6%). The
overall number of cystic lesions detected varied between EUS and
MRI (8 vs. 16). Communication between cysts and the pancreatic
duct (PD) was more often reported by EUS than MRI (4 vs. 1).
Conclusion Based on these preliminary results, EUS and MRI seem
complementary techniques to detect (pre)malignant lesions in indi-
viduals at high-risk for developing PC. MRI detected more cystic
lesions, but EUS detected communication between cyst and PD more
often. The latter is valuable information since it differentiates a
simple cyst from a side-branch IPMN. EUS detected one adenocar-
cinoma in these 33 patients, which was missed by MRI.
In familial adenomatous polyposis: multiple targeted
endoscopic biopsies are accurate in assessing severity
of duodenal adenomatosis
Musa Drini1, Anthony Speer2, Chris Dow3, Prithi Bhathal4,
Neil Collier5, Finlay Macrae1
1Colorectal Medicine and Genetics, 2Gastroenterology Department,3Anatomical Pathology, 5Department of Surgery, The Royal
Melbourne Hospital, Parkville Victoria 3050, Australia; 4Melbourne
Pathology, Collingwood, Victoria 3066, Australia
Background and study aims Duodenal polyps are common in FAP.
The cumulative risk for development of duodenal cancer is reported
to be 4.5% by the age of 57 year (95% CI 0.1–8.9%). Most upper
gastrointestinal screening protocols are based on Spigelman’s clas-
sification. However the published experience on surveillance of
duodenal adenomatosis in FAP, suggests that carcinomas identified in
screening are advanced and often not curable.
Patients and methods Review of FAP surveillance database between
January 1999 and November 2008 is presented.
Our surveillance protocol included multiple targeted endoscopic
biopsies of selected lesions. Upper Gastrointestinal endoscopy is
performed with side viewing instrument. Endoscopic findings were
classified as: macroscopically normal or small polyps, benign mac-
roscopically, suspicious for carcinoma macroscopically or confirmed
carcinoma.
Results Among 67 patients, 11 underwent surgical resection. Pan-
creas-preserving duodenectomy (PPD) was performed in four patients
(five procedures), and Whipple’s operation in seven patients. The
average size of polyps was 43 mm (range 17–65 mm), and the
average number of targeted endoscopic biopsies per lesion was 7.5
(range 5–10). The section of surgical specimens most representative
of the surface epithelium was evaluated for heterogeneity and from
this we calculated the approximate percentage of dysplasia/carcinoma
on the surface of the lesion, which would be amenable to endoscopic
biopsy. Two intramucosal carcinomas were diagnosed on endoscopic
biopsies, each understaged compared with the subsequent surgical
specimen. All carcinomas identified were resectable with no evidence
of local spread or distant metastasis. There was one postoperative
death a patient with significant co morbidities, but no cancer related
deaths.
Conclusion Our screening program identified two early carcinomas
and both underwent curative resections. Histology of resected polyps
shows considerable heterogeneity of dysplasia and or carcinoma
throughout polyps. Endoscopic biopsies understage some lesions.
High-risk lesions are best identified in a screening program by mul-
tiple targeted endoscopic biopsies.
What happens when you remove the duodenum from
a patient without a colon? Is bowel function or quality
of life impacted?
Yehuda Kariv, R. Kavir, A. da Luz Moreira, R. Mackay, Z. Kutalyli,
R. M. Walsh, J. Church
Cleveland Clinic Foundation, Cleveland, Ohio, USA
Background Patients with familial adenomatous polyposis (FAP)
and Spigelman IV duodenal adenomas need surgery to minimize
cancer risk. Options for duodenectomy include pancreas sparing
duodenectomy (PSD) and pancreaticoduodenectomy (PD). We studied
Abstracts 715
123
the influence of duodenal surgery and its specific types on functional
outcome and QOL in colectomized FAP patients.
Methods FAP patients who had undergone colon resection were
identified in a prospectively maintained polyposis registry. Long term
([1 year follow up from last surgery) function and QOL data were
obtained using a periodic questionnaire and completed by a telephone
survey. Outcomes in patients who had both colonic and duodenal
surgery (CDS) were compared to patients who underwent colonic
surgery without duodenal surgery (CS). CDS patients’ data was also
analyzed according to the specific type of colonic or duodenal
surgery.
Results 51 FAP patients underwent CDS. 9 patients died during
follow up (median, 192 (range 43–540) months from first surgery, 64
(range 19–250) months from last surgery). Long term functional and
QOL data were available in 33 CDS and compared to 249 CS patients
(Table). Median bowel movements per 24 h were 6 (CDS) and 5
(CS). Median quality of life (CCF Global Score) was 9/10 (CDS) and
9/10 (CS). Higher rates of diet (45% vs. 27%), social (19% vs. 5%),
sexual (17% vs. 5%) and work restrictions (21% vs. 9%) were
reported by CDS patients. CDS had lower quality of health scores.
CDS patients with an ileoanal pouch had more frequent bowel
movements but were similar in other functional and QOL parameters.
Outcomes were comparable between different types of duodenal
surgery.
Conclusions FAP patients needing duodenectomy can be reassured
that quality of life will be maintained.
Prophylactic pancreaticoduodenectomy for advanced
duodenal adenomatosis in familial adenomatous
polyposis
J. Skipworth1, C. Morkane1, N. West3, M. Deheragoda2, D. Raptis1,
C. Imber1, S. Olde-Damink1, M. Malago1, R. Phillips3, S. Clark3,
A. Shankar1
1Department of Hepatopancreaticobiliary Surgery, University College
London Hospital NHS Trust, London; 2Department of Pathology,
University College London Hospital NHS Trust, London;3The Polyposis Registry, St. Mark’s Hospital, Harrow
Background Patients with familial adenomatous polyposis (FAP)
develop duodenal polyps that may progress to malignancy, via the
adenoma-carcinoma sequence. In 2002, this centre reported data on
16 FAP patients who underwent prophylactic duodenal resection for
duodenal polyps. We now present the extended results, representing
the largest series of pancreaticoduodenectomy for FAP reported.
Methods We performed a retrospective case-notes review of all
patients undergoing prophylactic duodenal resection for advanced
duodenal adenomatosis, in a single centre. Data collected included
demographics, resection type, Spigelman staging, morbidity and
mortality.
Results 40 (24F:16M) FAP patients underwent prophylactic resec-
tion for advanced duodenal adenomatosis (Spigelman stage III/IV or
cancer) between January 1994 and November 2008. Data for 8 patients
were incomplete. Median patient age was 48 years and median hospital
stay 27 days. Available data revealed that 36 pancreaticoduodenec-
tomies and 1 local excision + cholecystectomy were performed. Peri-
operative mortality was 2 (5%): both patients dying from multi-organ
failure following re-operation for haemorrhage. Six further patients
have subsequently died, four from metastatic disease, one following a
cerebrovascular accident and data is missing in one case. Complica-
tions occurred in 24 (60%) patients. Early complications included six
anastomotic leaks, seven post-operative haemorrhages (three necessi-
tating redo laparotomy, one requiring gastroduodenal artery
embolisation and one needing embolisation of a jejunal vessel), three
enterocutaneous fistulae, one pulmonary embolus, one lymphatic leak
and 11 post-operative collections. Late complications included pan-
creatic insufficiency/steatorrhoea in 13 patients, diabetes in one and
delayed gastric emptying in four patients.
Postoperative histology revealed three (8%) pre-operatively
undetected ampullary/duodenal cancers—two of these patients have
subsequently died.
Conclusion Prophylactic pancreaticoduodenectomy is associated
with significant morbidity and mortality; however, FAP patients with
advanced duodenal polyposis have a significant risk of malignant
transformation and some have undetected malignancy. Thus, the
identification of FAP patients requiring surgical intervention for high-
risk of duodenal malignant transformation remains challenging.
Is there a genotype–phenotype correlation for ileal
pouch polyposis in patients with familial adenomatous
polyposis?
Alexander C. von Roon, Olivia C. Will, Ripple F. Man, Kay F. Neale,
R. John Nicholls, Robin K. S. Phillips, Paris P. Tekkis,
Susan K. Clark
The Polyposis Registry & Department of Surgery, St Mark’s Hospital
and Department of Biosurgery and Surgical Technology, Imperial
College, Harrow, Middlesex, UK
Aim To examine whether in patients with familial adenomatous
polyposis (FAP) who have undergone ileal pouch anal anastomosis
(IPAA), the severity of pouch polyposis is associated with APC codon
1309 mutation or number of intact 20 amino acid beta-catenin binding
and degradation sites (20aa repeats) on the mutant allele.
Methods All pouch endoscopy reports for patients with FAP
attending for annual surveillance after IPAA were reviewed. Only
patients with more than 10 years follow-up after IPAA and a known
APC mutation were included. The maximum incidence of pouch body
neoplasms and mutation status were recorded. Significance was
assessed with the Gamma statistic.
Results Of 206 patients who underwent IPAA, 54 met the inclusion
criteria. Fifteen patients had a codon 1309 mutation, 39 had other
mutations. A larger proportion of patients with a codon 1309 mutation
remained polyp-free (10/15 [67%] vs. 9/39 [23%]), a similar pro-
portion had 1–10 pouch polyps (4/15 [27%] vs. 11/39 [28%]), and a
lower proportion had over 10 pouch polyps (1/15 [7%] vs. 19/39
[49%]) after 10 years (p = 0.004). Of the 54 patients, 27 had muta-
tions resulting in no 20aa repeats, 19 had one repeat, 6 had 2 repeats
and 2 had 3 repeats. When compared with patients who had no 20aa
repeat, a larger proportion of those with one repeat remained polyp-
free (10/19 [53%] vs. 6/27 [22%]), a similar proportion had 1–10
polyps (7/19 [37%] vs. 6/27 [22%]), and a lower proportion had more
than 10 polyps (2/19 [11%] vs. 15/27 [56%]) after 10 years
(p = 0.025).
Conclusion The APC mutation at codon 1309 and mutations
resulting in a single 20aa repeat on the mutant allele appear to be
inversely correlated with the severity of pouch polyposis. This finding
contrasts with the genotype–phenotype correlation observed in colo-
nic polyposis. Larger studies are required to confirm these findings.
716 Abstracts
123
Children with familial adenomatous polyposis who
underwent early colectomy: psychological, quality
of life and pouch outcome
C. Durno, K. Butler, T. Berk, N. Alingary, M. J. Esplen
Dr. Zane Cohen Digestive Diseases Research Center, Familial
Gastrointestinal Cancer Registry and Department of Surgery, Mount
Sinai Hospital, Division of Gastroenterology, Hepatology and
Nutrition, Department of Paediatrics, Hospital for Sick Children,
University of Toronto, Toronto, Canada
Background The impact of ileal pouch-anal anastomosis on quality of
life in adolescents with FAP is favorable. There is a small group of
children with FAP who develop polyps at a younger age and have a
severe polyposis phenotype requiring earlier colectomy. The literature
does not focus on this very young subgroup. Patients who undergo early
colectomy may have a poor functional outcome and quality of life.
Aim To investigate functional outcome and quality of life in
patients with FAP who had colectomy \14 years of age.
Methods A cross-sectional quantitative survey was used to assess
patients with FAP recruited through a FAP Registry. Standardized
instruments included: psychosocial functioning, quality of life and
functional outcomes.
Results Among 1337 patients with FAP from 409 kindreds 59 (4%)
patients underwent colectomy at\14 years of age. Response rate was
84% (32/38). The mean age at the time of colectomy was 12 years
(SD 2), with a current mean age of 24 years (SD 8.5). Time since
colectomy was 12 years (SD 8.4, range 1–37 years). Bowel function:
78% of patients ‘‘rarely or never’’ have restrictions at work/school
and 60% are ‘‘always to sometimes’’ embarrassed. Patients currently
\18 years of age (9/32) have more restrictions. For the majority
psychosocial functioning was in normal ranges. Self and body esteem
were within normal ranges. Having a family member with FAP or
having lost a family member to FAP (11/32) did not affect quality of
life, self-esteem, or psychological outcome. Patients understand that
their risk of bowel cancer is still increased compared to the general
population. The majority of patients (n = 24, 75%) have had sur-
veillance endoscopy within the last 2 years.
Conclusions Patients who had early colectomies are functioning
well psychologically and with bowel function. Health-related quality
of life and compliance with endoscopic surveillance is very good. A
subgroup who are currently under 18 years of age demonstrate
emotional issues and difficulty adapting. There are some restrictions
due to bowel and psychological symptoms. This subgroup would
benefit with added psychological interventions to enhance coping.
MyFAP: An internet-based psychosocial intervention
for adolescents and young adults with FAP
Susan K. Peterson, Martha Askins, Alex Prokhorov, Devki Saraiya,
Thuy Vu, Miguel Rodriguez-Bigas, Patrick Lynch
The University of Texas M. D. Anderson Cancer Center, Houston,
Texas, USA
Adolescents and young adults (AYAs) with familial adenomatous
polyposis (FAP) face unique medical and psychosocial demands
related to genetic counseling and testing, routine colorectal screening,
and preventive colectomy. There are scant informational and sup-
portive resources available for this population. The Internet is a
promising medium for the delivery of psychosocial interventions to
persons with rare conditions, and Internet use by young persons is
nearly universal. We describe the development and initial evaluation
of MyFAP, an Internet-based, multimedia psychosocial intervention
for AYAs with FAP. The goal of MyFAP is to facilitate self-man-
agement and coping with the medical and psychological issues faced
by young persons with FAP. Grounded in Social Cognitive Theory
(SCT), MyFAP includes the following components: medical and
genetic aspects of FAP; self-management (e.g., screening, self-care
issues); emotional concerns and support; communication with peers,
family and health care providers; and, preventive surgery. Consistent
with SCT, the intervention includes multiple elements to facilitate the
adoption of coping and problem-solving skills through cognitive-
behavioral activities and interactive multimedia features. MyFAP
includes a social networking component to promote social interaction
and support with peers. Twenty-three AYAs age 13–24 years com-
pleted an initial evaluation of the intervention. The mean correct score
on a baseline measure of FAP-related knowledge was 51%, indicating
the need for improved education about clinical aspects of FAP.
Greater than 90% of users rated the highest level of satisfaction
with MyFAP’s attractiveness, control, efficiency, helpfulness, and
learn ability, using a standardized measure. Findings showed that an
Internet-based psychosocial intervention is a feasible and acceptable
method for addressing the informational and supportive needs of
AYAs with FAP.
Compliance with endoscopic surveillance advice
for familial adenomatous polyposis (FAP):
room for improvement
Kirsten F. L. Douma, Eveline M. A. Bleiker, Neil K. Aaronson,
Annemieke Cats, Miranda A. Gerritsma, Chad M. Gundy,
Hans F. A. Vasen
The Netherlands Cancer Institute, Antoni van Leeuwenhoek hospital,
Amsterdam, The Netherlands
Background & Aims Familial adenomatous polyposis (FAP) is
characterized by the development of hundreds of adenomas in the
colorectum that, without surgery, lead to colorectal cancer. The purpose
of the study was to assess compliance with endoscopic surveillance
advice before as well as after (prophylactic) surgery for FAP.
Methods In this nationwide, cross-sectional study, individuals from
families at high risk for FAP registered with the Netherlands Foun-
dation for the Detection of Hereditary Tumours were invited to
complete a questionnaire on psychosocial issues and endoscopic
screening experiences. Compliance data were derived from medical
records and via self-report.
Results A total of 328 individuals were eligible for the study, of
whom 85 were at risk for FAP, 108 had an intact rectum after a
colectomy with ileorectal anastomosis (IRA), and 135 had a pouch
following a proctocolectomy with ileoanal anastomosis (IPAA).
Based on medical record data, 20% of the at-risk group and 6% of the
IRA group were found to be less than fully compliant with surveil-
lance advice. Under-compliance in the at-risk group was associated
significantly with perceived self-efficacy, use of sedatives during
surveillance, pain after surveillance and low perceived benefits of
surveillance (p \ .05).
Conclusions One-fifth of individuals at-risk for FAP are under-
compliant with screening advice. Low self-efficacy, non-use of sed-
atives during surveillance and pain after surveillance are negatively
associated with compliance behavior. We recommend that sedatives
be routinely offered to individuals undergoing colorectal cancer sur-
veillance for FAP and that adequate pain medication or spasmolytica
be provided after endoscopy.
Abstracts 717
123
Familial adenomatous polyposis (FAP): attitudes
towards genetic testing in childhood and reproductive
decision-making
Kirsten F. L. Douma, Neil K. Aaronson, Hans F. A. Vasen,
Senno Verhoef, Chad M. Gundy, Eveline M. A. Bleiker
The Netherlands Cancer Institute—Antoni van Leeuwenhoek
Hospital, Amsterdam, The Netherlands
Purpose Childhood DNA testing, prenatal diagnosis (PND) and pre-
implantation genetic diagnosis (PGD) are available for most heredi-
tary cancer susceptibility syndromes. However, the use of PND and
PGD for these syndromes is controversial. The purpose of this study
is to investigate attitudes towards, and experiences with, childhood
DNA testing, PND and PGD among members of families at high risk
for familial adenomatous polyposis (FAP).
Methods In this nationwide cross-sectional study, individuals from
families at high risk for FAP, were invited to participate. Question-
naire data were collected on attitudes towards and experiences with
childhood testing, PND and PGD, and on a range of sociodemo-
graphic, clinical and psychosocial variables.
Results 525 FAP-family members participated (response rate =
64%). Forty percent of FAP-patients expressed that the disease has
influenced their desire to have children. Only a small proportion
(15%) considered termination of pregnancy for FAP acceptable.
Approximately 30% of individuals with a FAP-diagnosis and their
partners had a positive attitude towards PND and PGD. A positive
attitude towards PND and PGD was associated with higher levels of
guilt and a positive attitude towards termination of pregnancy. Five
individuals reported direct personal experience with PND and one
with PGD. Ninety-three individuals with a FAP-diagnosis and 43
partners had children who were minors (\18 years) during the DNA
testing procedure. Most of these parents (82%) were satisfied with the
procedure. One-third of the individuals wanted DNA testing for their
children before age 12.
Conclusion FAP family members’ experiences with childhood DNA
testing are predominantly positive. Approximately one-third of those
with a FAP diagnosis have a positive attitude towards PND and PGD.
Few, however, have had direct experience with these procedures.
Extracolonic tumour spectrum and incidence in 276
patients affected by MUTYH-associated polyposis
(MAP)
Stefan Aretz, Daria Christian, Christoph Engel, Maartje Nielsen,
Natalie Jones, Astrid Kaufmann, Verena Steinke, Hans F. Vasen,
Peter Propping, Frederik J. Hes, Julian R. Sampson
Stefanie Vogt Institute of Human Genetics, Bonn, Germany
Background To date, no systematic evaluation of extracolonic MAP
manifestations has been reported.
Methods In a collaborative multicentre European study a large
cohort of MAP patients (276 cases from 181 apparently unrelated
families) was recruited. Based on medical records and anamnestic
information the extracolonic tumour spectrum and incidence were
evaluated to assess cumulative lifetime risks and compared with
general population rates to obtain standardized incidence ratios (SIRs).
Results The median age at evaluation was 54 years. Duodenal
polyposis occurred in 17%; the relative risk of duodenal cancer was
very high (SIR 130; 95% CI 16–470), while the lifetime risk was
3.6%. Regarding extraintestinal tumours we found a low to moderate
but significant increase in the incidence of breast cancer (SIR 3.0;
95% CI 1.5–5.4; mean age at diagnosis 60 years), bladder carcinomas
(SIR 7.23; 95% CI 1.97–18.5; mean age at diagnosis 59 years), and
skin cancer (SIR 2.8; 95% CI 1.5–4.8; mean age at diagnosis
52 years), other malignant lesions were not increased significantly.
The incidence of extraintestinal malignancies as a whole was almost
doubled (SIR 1.8; 95% CI 1.3–2.4), the lifetime risk was 40% (95%
CI 24–55%). Interestingly, sebaceous gland tumours (SGT), that are a
characteristic of Muir–Torre syndrome, occurred in five patients. No
genotype–phenotype correlation was identified.
Conclusions The relative risks of a few cancers and of extraintestinal
malignancies as a whole were increased, however, no predominant
lesion was observed. The spectrum of cancers and their advanced age
at onset do not suggest that specific surveillance recommendations
other than frequent gastrointestinal endoscopies can be made at
present. Although not significant, a trend towards a slightly increased
frequency of gynaecological tumours (endometrial and ovarian can-
cer) and SGT point to a phenotypic overlap with Lynch syndrome.
SGTs might serve as diagnostic marker lesion in some cases.
The study was supported by the Deutsche Krebshilfe, the Dutch
Digestive Diseases Foundation, and the Wales Gene Park and Cancer
Research Wales.
Analysis of MUTYH genotypes and colorectal
phenotypes in patients with MUTYH associated
polyposis
Maartje Nielsen, Mirjam C. Joerink-van de Beld, Natalie Jones,
Stefanie Vogt, Carli M. Tops, Hans F. A. Vasen, Julian R. Sampson,
Stefan Aretz, Frederik J. Hes
Department of Clinical Genetics, Leiden University Medical Center
(MN, MCJB, CMT, FJH), Institute of Medical Genetics, School
of Medicine, Cardiff University (NJ, JRS), Institute of Human
Genetics, University of Bonn (SV, SA), Department
of Gastroenterology & Medical Oncology, Leiden University
Medical Center (HFAV)
Background & Aims Functional studies have demonstrated signifi-
cant differences in base recognition and glycosylase activity between
various MUTYH mutations, notably for the two mutations most fre-
quently reported in MAP patients: Y179C and G396D (previously
annotated as Y165C and G382D). Our goal was to establish correlations
between genotypes and colorectal phenotype of patients with MAP.
Methods In this multicenter study, we analyzed genotype and phe-
notype data from 257 MAP patients. Data included age at presentation
of MAP, polyp count, the occurrence, location and age at presentation
of CRC.
Results Patients with a homozygous G396D mutation or compound
heterozygous G396D/Y179C mutations presented later with MAP and
had a significantly lower hazard of developing CRC than patients with
a homozygous Y179C mutation (P \ 0.001). The mean ages of CRC
diagnosis in patients were 58 years (homozygous G396D) and
52 years (compound heterozygous G396D/Y179C) versus 46 years
(homozygous Y179C; P = 0.001, linear regression).
Conclusions Our study identified the phenotypic effects of Y179C
as relatively severe and of G396D as relatively mild. These clinical
data are in accord with findings from in vitro functional assays.
Genotypic stratification may become useful in the development of
guidelines for counseling, surveillance and management of families
with MAP.
718 Abstracts
123
1. Harris M (2008) Why all young bowel cancer patients should be
screened for Lynch syndrome. ANZ J Surg 78:531–532.
Renal cancer as part of the phenotype
of MYH-associated polyposis; the evidence
gets stronger
Lisa LaGuardia, Margaret O’Malley, Carol Burke, James Church
Department of Colorectal Surgery, The Sanford R. Weiss, M. D.
Center for Hereditary Colorectal Neoplasia, Digestive Disease
Institute, Cleveland Clinic, Cleveland Ohio
Background The full phenotype of MYH-Associated Polyposis
(MAP) is still not known as the syndrome is relatively new and the
number of affected families relatively sparse. The syndrome often
seems to mimic attenuated FAP except that the pattern of inheritance
is recessive rather than dominant. In 2006 a preliminary study of the
tumor spectrum that was seen in 6 MAP families suggested that renal
cancer may be found to excess. Now we have 7 additional MAP
families with information that more strongly implicates renal cancer
as part of the syndrome.
Methods The families of thirteen probands with MAP were char-
acterized by extensive pedigree building. Data was entered into the
Polyposis registry database. Where available, pathology reports were
requested to document the reliability of the family history. All pro-
bands had biallelic mutations of MYH.
Results Relatives under the age of eighteen and the spouses of
at-risk relatives are excluded. There were 247 at-risk relatives in the
13 families. There were 67 affected (27%), some with multiple
tumors. The cancer spectrum is 23 patients had colon cancer out of 8
families, 6 had prostate cancer out of 3, 6 had renal cancers out of 6, 2
had uterine cancer out of 1, 2 had pancreatic cancers out of 1, 4 had
breast cancers out of 4, 1 had bone cancer out of 1, 1 had brain cancer
out of 1, 5 had lung cancers out of 5, and 2 had leukemia out of 2.
Discussion The colorectal phenotype of these MAP families is as
expected. 13.3% of relatives had adenomas although less than half of
the relatives had screening colonoscopy. Colon cancers were found in
23 individuals, usually with an older age of onset. The spectrum of
extracolonic cancers seen in these 13 families is unusual. Renal
cancer occurred in six different families. There were four cases of
prostate cancer in one family and two with pancreatic cancer in
another 78% of patients with a cancer had at least one Y165C
mutation; 62% had at least one G382D mutation.
Conclusion The phenotype of MAP is evolving. Detailed studies on
phenotype in much large numbers of families are needed. Renal
cancer was present 46% of these families and renal ultrasound is
recommended as part of the surveillance program.
Changing causes of death in familial adenomatous
polyposis: signs of progress but more work to do
Lisa LaGuardia, Margaret O’Malley, James Church, Carol Burke,
Matthew Kalady
The Sanford R. Weiss, MD, Center for Hereditary Colorectal
Neoplasia, Digestive Disease Institute, Cleveland Clinic, Cleveland,
Ohio
Introduction Patients with FAP are at risk of dying from multiple
benign and malignant tumors, from surgical complications, comorbid
diseases and the rigors of life in the twenty-first century. Our last
study on why patients with FAP die was in 1990. We have analyzed
causes of death since then and compared the two groups to see if the
significant advances in medicine and technology are reflected in
different patterns.
Methods Causes of death were extracted from the 1990 study via
the manuscript. Causes of death since then were determined from the
registry database and confirmed by chart review.
Results In 1990 there were 178 FAP families in the registry. There
are now 761 families. 212 patients have died (0.28 deaths perfamily),
110 before 1990 (0.62 deaths perfamily) and 102 since 1990 (0.17
deaths per family). Death from colorectal cancer before 1990 were 64
(58.2%), and after 1990 were 41 (40.2%). Deaths from desmoid
disease were 12 (10.9%), and 9 (8.8%), from periampullary cancer
were 9 (8.2%), and 4 (3.9%), from brain cancer were 8 (7.3%), and 2
(2.0%), Perioperative deaths were 5 (4.5%), and 3 (2.6%). Accidental
deaths were 3 (2.7%), and 0 and death from other causes were 9
(8.2%), and 24 (23.5%). Unknown 0, and 19 (18.6%). Overall death
from cancer (excluding desmoids) was 85 (77.3%) before 1990 and
57 (55.9%) since 1990. ‘‘Other’’ deaths include cancers of the thyroid,
stomach, esophagus, pancreas, breast, ovary and lung. There were
also 2 suicides and one death from pancreatitis. Overall there have
been fewer deaths per family since 1990. Deaths from colorectal and
periampullary/duodenal cancer have declined (even if the ‘‘unknown’’
category is excluded) while those for desmoid remain constant. Peri-
operative deaths are fewer.
Conclusion Improvements in education and more access to genetic
testing, screening and surgical techniques should further reduce the
death rate from FAP. There is scope for doing so.
Interobserver variability in the interpretation
of immunohistochemical mismatch repair protein
staining
Louise Klarskov1, Susanne Holck1, Karina Ronlund2,
Jan Lindebjerg2, Jacob Elebro3, Britta Halvarsson4,
Inge Bernstein5, Jenny von Salomee6, Mef Nilbert7
1Department of Pathology, Faculty of Health Sciences, Copenhagen
University, Hvidovre Hospital, Copenhagen, Denmark; 2Department
of Pathology, Vejle Hospital, Vejle, Denmark; 3Department of
Pathology, Lund University Hospital, Lund, Sweden; 4Department
of Pathology, HNPCC-register, Department of Gastroenterology,
Copenhagen; 5University, Hvidovre Hospital, Copenhagen, Denmark
Helsingborg Hospital, Helsingborg, Sweden; 6Department of Clinical
Genetics, Stockholm University, Karolinska Hospital, Stockholm,
Sweden; 7Clinical Research Centre, Faculty of Health Sciences,
Copenhagen University, Hvidovre Hospital, Copenhagen, Denmark
Introduction Immunohistochemical staining for mismatch repair
(MMR) proteins is a widely implemented diagnostic tool in the work
up of suspected hereditary colorectal cancer, but quality assurance
studies are scarce. In order to validate routine staining with respect to
interobserver variability, we reviewed MMR protein immunostainings
from 225 colorectal cancers.
Materials and methods All tumors were stained as part of routine
examination linked to genetic counselling for suspected hereditary
colorectal cancer. Immunohistochemical stainings for MLH1, PMS2,
MSH2 and MSH6 were reviewed in a blinded fashion. In order to test
the impact of experience in immunohistochemical evaluation, 3
pathologists specialized in gastrointestinal diagnostics and 2 residents
in pathology evaluated all samples. Stainings were evaluated as
normal, completely lost, weak or non-evaluable. Consensus was
Abstracts 719
123
defined as all five observers agreeing on one of the four predeter-
mined staining patterns.
Results Consensus was reached for MLH1 in 52%, PMS2 in 60%,
MSH2 in 81%, and MSH6 in 43% of the stainings. Related kappa-
values of interobserver agreement, two by two, varied between
0.35–0.81, 0.45–0.91, 0.61–0.95, and 0.23–0.77, respectively. Kappa-
values among the 3 specialists did not differ from the values between
the 2 residents. Discrepancy was predominantly related to cases
judged weak instead of normal or completely lost by at least one of
the observers, whereas discrepancy between normal and completely
lost occurred in 2–5% of the stainings.
Discussion Improved HNPCC diagnostics is crucial since it allows
identification of high-risk individuals who should be offered partici-
pation in surveillance programmes. Herein standardization and
validation of MMR protein immunostaining evaluation is essential.
Our findings suggest that differences in interpretations of the MMR
protein immunohistochemical stainings are common, which under-
scores the need for quality assurance programmes.
Narrow-band imaging improves the detection of polyps
in patients with hyperplastic polyposis syndrome:
a prospective randomized study
K. S. Boparai, F. J. C. van den Broek, S. van Eeden, P. Fockens,
E. Dekker
Academic Medical Centre, Amsterdam, The Netherlands
Background Hyperplastic polyposis syndrome (HPS) is associated
with colorectal cancer (CRC). HPS patients receive endoscopic sur-
veillance to prevent malignant progression of polyps. Endoscopic
detection and removal of potentially premalignant sessile serrated
adenomas (SSAs) and conventional adenomas may be an important
step in preventing CRC development in HPS. However, polyps in
HPS can be difficult to detect due to their unremarkable color and flat
shape.
Objective To prospectively compare the value of narrow-band
imaging (NBI) and high-resolution white light endoscopy (WLE) for
the detection of polyps in HPS and to assess the value of NBI for the
differentiation of these polyps.
Patients and interventions During surveillance endoscopy in 22 HPS
patients, each colonic segment was inspected twice, once with WLE
and once with NBI, in random order by one experienced endoscopist.
Of all detected polyps the size, shape and location was assessed. Kudo
pit-pattern analysis was performed in all polyps.
Main outcome measurements The sensitivity of WLE and NBI for
the detection of polyps was assessed. The diagnostic accuracy of NBI
in differentiating detected polyps was determined by using histology
as a gold standard.
Results A total of 116 HPs, 42 SSAs and 24 adenomas (range:
2–20 mm) were detected. Polyps were classified as flat (60%), sessile
(38%) and pedunculated (2%).
The sensitivities of WLE and NBI for overall polyp detection were
64 and 90% respectively (p \ 0.001). For flat polyps these were 51
and 87% (p \ 0.001) and for sessile/pedunculated polyps 81 and 96%
(ns). The sensitivities of WLE and NBI for HPs was 67 and 90%
respectively (p = 0.017). For SSAs this was 38 and 86% (p = 0.003)
and for adenomas 70 and 100% (ns). The accuracy of NBI for dis-
criminating SSAs from HPs was 63% and for differentiating
adenomas from HPs this was 75%.
Conclusion NBI significantly improves the detection rate of SSAs
and predominant flat polyps in HPS. Therefore, NBI seems of clinical
value for the endoscopic management of HPS patients.
Family history of colorectal cancer is inversely related
to polyp count in individuals with multiple serrated
polyps
Joanne Young, Daniel D. Buchanan, Kevin Sweet, Musa Drini, Mark
A. Jenkins, Aung Ko Win, Michael Gattas, Michael D. Walsh, Diane
McKeone, Aedan Roberts, Mark Clendenning, Rhiannon Walters,
Sven Arnold, Alasdair Young, Heather Hampel, John L. Hopper, Jack
Goldblatt, Jill George, Graeme K. Suthers, Kerry Phillips, Graeme
P. Young, Elizabeth Chow, Susan Parry, Kathy Tucker, Amanda
Muir, Michael Field, Sian Greening, Steven Gallinger, Jane Green,
Michael O. Woods, Renee Spaetgens, Albert de la Chapelle, Finlay
Macrae, Jeremy R. Jass
Familial Cancer Laboratory, Queensland Institute of Medical
Research, Herston, Australia
Hyperplastic polyposis (HPS) is a colonic polyposis condition of
unknown aetiology. An association with an increased risk of colo-
rectal cancer (CRC) has previously been established. The purpose of
this study was to examine the spectrum of phenotypic variation in
HPS.
One hundred and twenty-six patients with HPS were recruited to
the study. Patients were assigned to three categories based on polyp
numbers which were extracted from histology and colonoscopy
reports. Ethnicity of paternal and maternal lines was self-reported.
Family history of colorectal cancer data were derived from pedigrees.
All patients were screened for the two most common mutations in
MUTYH. One hundred and fourteen of 120 (95%) HPS probands
were of white, northern European ethnicity.
One of 126 patients (\1%) was homozygous for the Y165C var-
iant in the MUTYH gene. The average minimum reported polyp
number was 39; 28, 59 and 13% of the participants were in the polyp
category 1 (5–20 polyps), category 2 (21–70 polyps) and category 3
([70 polyps), respectively.
CRC was identified in 49 of 119 patients (41%) and 28% of these
patients had multiple CRC. CRC was significantly associated with the
presence of adenomas (P = 0.03). Overall, 59% of probands had at
least one-first-degree relative affected with CRC. Family history of
CRC (especially a first-degree relative with CRC) was inversely
associated with polyp number categories (P = 0.03). In addition,
males were more likely to have higher numbers of serrated polyps
(P = 0.04).
We conclude that HPS is associated with an increased personal
risk of CRC, and higher polyp numbers in males. The inverse rela-
tionship between polyp numbers and family history of CRC suggests
heterogeneous modes of inheritance.
Serological markers in evaluation of individual risk
of gastric cancer among mutation carriers in Lynch
syndrome
39 Mecklin, A. Ristimaki, P. Sipponen, K. Nuorva, V. Karja,
S. Sarna, L. Renkonen-Sinisalo, M. Aarnio, M. Heikkinen,
K. Pylvanainen, H. J. Jarvinen
Jyvaskyla Central Hospital, Jyvaskyla, Finland
720 Abstracts
123
Gastric cancer (GC) is the second or third most common cancer in
Lynch syndrome (LS), but its mortality often exceeds the mortality of
colorectal or endometrial cancers. A chronic, active infection by high
virulence H. pylori strains can transform normal gastric mucosa into
atrophic gastritis. This type of response to the chronic infection may
eventually lead to dysplastic changes and transformation to GC. Over
90% of GCs in LS seems to be histologically of intestinal type (IT).
Therefore, H. pylori infection and atrophic gastritis can be considered
as markers of increased risk of GC in individuals with LS and
indicative for interventions.
Gastroscopy has been considered the only screening method
available, but its cost-benefit is low especially in most western
countries. Our purpose was to find a cost-benefit non-endoscopic
alternative to identify Lynch syndrome mutation carriers at risk for
gastric cancer.
Experimental design 89 Lynch syndrome mutation carriers over
50 years underwent upper-GI-endoscopy with biopsies and blood test
panel including pepsinogen I and II, fasting gastrin-17 and immu-
noglobin G antibodies to H. pylori.
Results Normal gastric mucosa was diagnosed in 71.6–77.3%,
superficial gastritis in 21.6–18.2% and atrophic gastritis in 6.8–4.5%
by serology and histology, respectively. The sensitivity of serology to
identify atrophic gastritis was 0.750 (95% CI 0.301–0.954) and
specificity 0.988 (95% CI 0.936–0.988). Positive predictive value was
0.750 (95% CI 0.301–0.954) and negative predictive value 0.988
(95% CI 0.936–0.998). The serological test panel overestimated
normal mucosa into superficial gastritis in four and atrophy in one
cases.
Conclusion Serological biomarkers identify reliably atrophic gas-
tritis and they can be used as non-endoscopic screening method in
evaluating individual risk for gastric cancer and need for endoscopic
surveillance in Lynch syndrome in countries with a low general
incidence for gastric cancer.
Natural history of Amsterdam patients following
colorectal cancer resection
Matthew F. Kalady, James Church, Jon Vogel, Ellen McGannon,
Susan Fay, Elena Manilich, Lori Arroyo, Kathy Toderick,
Janet Shenal
Department of Colorectal Surgery, Sanford R. Weiss Center
for Hereditary Colorectal Neoplasia, Cleveland Clinic, Cleveland,
OH, USA
Introduction Colorectal cancer patients meeting Amsterdam criteria
are offered total rather than segmental surgical resection to reduce the
risk of metachronous colorectal malignancy. There is sparse natural
history information, however, for patients who undergo a segmental
colectomy. This study evaluates the natural history of surgically
treated Amsterdam criteria colorectal cancer patients.
Methods A single institution hereditary colorectal cancer database
was reviewed for all patients meeting Amsterdam I, II or-like criteria
or germline-confirmed Lynch patients who underwent surgical resec-
tion. Hereditary syndrome, patient demographics, type of surgery
performed, tumor characteristics and subsequent follow-up were
recorded. The primary endpoints for patients undergoing surgery less
than a total proctocolectomy were subsequent adenoma formation and
further resection for cancer.
Results 208 patients were included, 67 treated primarily at our
institution and 141 referred to our registry after undergoing index
resection. 49% were female and 173 cancers (83%) were right-
sided. The median age at index surgery was 51.9 years. 176
patients underwent a partial colectomy, 23 underwent a total or
subtotal colectomy with ileorectal or ileosigmoid anastomosis,
respectively, and 9 patients underwent a total proctocolectomy. 66
of 199 patients (33%) underwent subsequent polypectomy during
surveillance, with removal of 131 adenomas including 24 that were
[10 mm and 30 with tubulovillous or villous architecture. 25 of
176 (14%) who underwent a partial colectomy subsequently
developed a colorectal cancer requiring resection at mean time of
124 months from index surgery. The stages at second resection
were I-9, II-11, III-4.
Conclusions Amsterdam patients undergoing partial colectomy
have a significant rate of metachronous high-risk adenoma formation
and cancer development, some of which present at advanced stage.
Therefore, total colectomy is still advocated as the index surgery.
However, if circumstances result in a segmental colectomy, patients
should undergo timely surveillance protocols and intervention to
prevent future malignancies.
The outcome of longterm surveillance of Lynch
syndrome families in the Netherlands
H. F. A. Vasen1, J. Kleibeuker2, M. van Kouwen3, J. J. Koornstra2,
A. Cats4, E. Dekker5, A. M. J. Langers1, S. Sanduleanu6,
J.-W. Poley7, J. C. H. Hardwick1, W. H. de Vos tot Nederveen
Cappel1, A. E. van der Meulen-deJong1, F. N. Nagengast3,
The Dutch Lynch Syndrome Study Group8
Departments of Gastroenterology, University Medical Centre
of Leiden1, Groningen2, Nijmegen3, Netherlands Cancer Institute
Amsterdam4, AMC Amsterdam5, Maastricht6, Rotterdam7
and The Netherlands Foundation for the Detection of Hereditary
Tumours8
Background Carriers of an MMR gene mutation have a high risk of
developing colorectal cancer (CRC). Because of evidence of an
accelerated colorectal carcinogenesis in Lynch syndrome, the sur-
veillance protocol for these families has been changed in the mid
Nineties. Since then, the recommended interval between examina-
tions has been 1–2 years in stead of 2–3 years. The aim of the study
was (1) to evaluate the risk of developing CRC while under sur-
veillance and (2) to identify risk factors.
Patients and methods The database of the Registry was used. Only
carriers of an MMR mutation who had more than one surveillance
examination were selected. MMR gene carriers who underwent a
previous colectomy were excluded. The observation time was from
1-1-1995 until 1-1-2008. Endpoints of the study were CRC, death or
the last colonoscopy. Kaplan–Meyer analysis was used to calculate
the risk of CRC.
Results A total of 721 mutation carriers were included. The mean
follow up was 6.7 years. Thirty-three patients developed CRC. 85%
were at stage I or II. The cumulative risk of developing CRC was 5%
at 10 years of follow up. Male carriers had a (nonsignificant) higher
risk than female carriers. Carriers of an MLH1 or MSH2 mutation had
a significant higher risk than MSH6 carriers. There was no difference
in risk between surveillance at university medical centres and
peripheral hospitals.
Conclusions The risk of developing CRC under surveillance has
decreased since shortening of the surveillance interval. The risk was
highest in carriers of an MSH2 and MLH1-gene mutation. In these
groups annual colonoscopy and/or the use of chromoendoscopy may
be considered.
Abstracts 721
123
Modifiers of colorectal cancer risk in Lynch syndrome
J. T. Wijnen, R. M. Brohet, S. Jagmohan-Changur, R. van Eijk,
A. Middeldorp, C. M. Tops, M. van Puijenbroek, M. G. E. M.
Ausems, E. Gomez Garcıa, F. J. Hes, N. Hoogerbrugge, F. H. Menko,
T. A. M. van Os, R. H. Sijmons, S. Verhoef, A. Wagner, F. M.
Nagengast, J. H. Kleibeuker, P. Devilee, H. Morreau,
I. P. Tomlinson, R. S. Houlston, T. van Wezel, H. F. A. Vasen
Leiden University medical Center, Leiden, The Netherlands
Recent genome-wide association studies have identified common low
risk variants for colorectal cancer (CRC). To assess whether these
variants influence CRC risk in the Lynch syndrome (LS), we geno-
typed these variants in 675 proven MMR mutation carriers from 127
different LS families. We used univariate and multivariate analysis to
analyse the association between the presence of a risk variant and
CRC risk.
In our initial analysis of six variants on 8q24.21, 8q23.3, 10p14,
11q23.1, 15q13.3 and 18q21.1 we found a significant association
between CRC risk and rs16892766 (8q23.3) and rs3802842
(11q23.1). The C allele of rs16892766 was associated with an ele-
vated risk of CRC in a dose dependent fashion, with homozygosity for
CC conferring a 2.16-fold increased risk compared to the AA
homozygotes. For rs3802842 the increased risk of CRC associated
with the C-allele was only found among female carriers, while CRC
risk was substantially higher among homozygous (HR 3.08) than
among heterozygous carriers of the C-allele (HR 1.49). In an additive
model of both variants, the CRC risk was significantly associated with
the number of risk alleles (HR 1.60 for carriers of two or more risk
alleles compared to carriers of none or one risk allel). The observed
effects were stronger in female carriers than in male carriers. Results
on genotyping of eight additional low risk variants will be presented
at the meeting.
Conclusion So far we have identified two loci which are signifi-
cantly associated with CRC risk in Lynch syndrome families that may
be helpful to identify high risk individuals that require more intensive
surveillance.
Haemochromatosis HFE gene polymorphisms
as potential modifiers of hereditary nonpolyposis
colorectal cancer risk and onset age
R. J. Scott, Z. Shi, D. Johnstone, B. A. Talseth-Palmer, T. Evans,
A. D. Spigelman, C. Groombridge, E. A. Milward, J. K. Olynyk,
J. Suchy, G. Kurzawski, J. Lubinski
Hereditary nonpolyposis colorectal cancer (HNPCC) is characterised
by germline mutations in DNA mismatch repair genes, however
variation in disease expression suggests there are potential modifying
factors. Polymorphisms of the HFE gene, which cause the iron
overload disorder hereditary haemochromatosis, have been proposed
as potential risk factors for the development of colorectal cancer
(CRC).
To understand the relationship between HNPCC disease pheno-
type and polymorphisms of the HFE gene a total of 362 individuals
from Australia and Poland with confirmed causative MMR gene
mutations were genotyped for the HFE C282Y and H63D
polymorphisms.
A significantly increased risk of developing CRC was observed for
H63D homozygotes when compared to combined wild type homo-
zygotes and heterozygotes (hazard ratio = 2.93, p = 0.007).
Evidence for earlier CRC onset was also observed in H63D homo-
zygotes with a median age of onset 6 years earlier than wild type or
heterozygous participants (44 vs. 50 years of age). This effect was
significant by all tests used (log-rank test p = 0.026, Wilcoxon
p = 0.044, Tarone–Ware p = 0.035). No association was identified
for heterozygosity of either polymorphism and limitations on power
prevented investigation of C282Y homozygosity or compound
C282Y/H63D heterozygosity. In the Australian sample only, women
had a significantly reduced risk of developing CRC when compared to
men (hazard ratio = 0.58, p = 0.012) independent of HFE genotype
for either SNP.
In conclusion homozygosity for the HFE H63D polymorphism
appears to be a genetic modifier of disease expression in HNPCC.
Understanding the mechanisms by which HFE interrelates with
colorectal malignancies could lead to reduction of disease risk in
HNPCC.
Development and validation of a prediction model
to estimate gene-specific risk in Lynch syndrome
Fay Kastrinos1, Ewout W. Steyerberg2, Judith Balmana3, Rowena
Mercado4, Sapna Syngal5
1Herbert Irving Comprehensive Cancer Center, Columbia University
Medical Center, New York, New York, USA; 2Erasmus Medical
Center, Rotterdam, The Netherlands; 3Hospital Vall d’Hebron,
Universitat Autonoma de Barcelona, Spain; 4Dana-Farber Cancer
Institute, Boston, Massachusetts, USA; 5Dana-Farber Cancer
Institute, Brigham and Women s Hospital, Boston, Massachusetts,
USA
Background and aims Lynch Syndrome is caused by mutations in
the mismatch repair (MMR) system. Our aim was to expand a pre-
vious clinical model predicting the likelihood of finding a deleterious
gene mutation in at-risk patients incorporating MSH6 prediction and
gene-specific risk estimates.
Methods We analyzed an unreported cohort of 4538 unrelated
probands undergoing clinical genetic testing for MLH1, MSH2 and
MSH6 mutations at a commercial laboratory. Personal and family
history of cancer, cancer types and ages at diagnosis were compared
by gene mutation for probands and their first- and second-degree
relatives. A multivariable model using logistic regression was
developed to predict the likelihood of finding a MMR gene mutation
and provide a risk estimate for each individual gene. Validation was
by bootstrap resampling and independent validation in one-third of
the cohort.
Results Twelve percent (525/4538) of subjects had pathogenic
mutations (204 MLH1, 250 MSH2, 71 MSH6). Strong predictors of
MLH1 and MSH2 mutations included CRC history in probands (OR:
5.1, 4.5 for MLH1 and MSH2, respectively) and relatives (OR: 3.3 for
MLH1 and MSH2), whereas CRC was not as predictive of a MSH6
gene mutation. Endometrial cancer among probands was predictive of
MSH2 and MSH6 mutations (OR: 7.2 for both genes) with younger
age of diagnosis most notable for a MSH2 mutation. The model
discriminated well at external validation for each gene, with area
under receiver operating characteristic curve of 0.85 (95% CI 0.80–
0.89) for MLH1, 0.87 (95% CI 0.83–0.92) for MSH2, and 0.80 (95%
CI 0.68–0.92) for MSH6.
Conclusions From this large cohort of MMR gene mutation car-
riers, we expanded a clinical prediction model to evaluate an
individual’s gene-specific probability of being a mutation carrier in
MLH1, MSH2, and MSH6 genes. Our model’s capacity to estimate
mutation probability by affected gene offers healthcare professionals
ability to convey clinical information in an individualized quanti-
tative way.
722 Abstracts
123
Identification of novel genes involved in colorectal
cancer predisposition
Ramprasath Venkatachalam1, Marjolijn J. L. Ligtenberg1,2, Eveline J.
Kamping1, Eveline Hoenselaar1, Marsha Voorendt1, Heike Gorgens3,
Hans K. Schackert3, Ad Geurts van Kessel1, Nicoline Hoogerbrugge1,
Roland P. Kuiper1
1Departments of Human Genetics,2Pathology, Radboud University
Nijmegen Medical Centre, Nijmegen Centre for Molecular Life
Sciences, Nijmegen, The Netherlands; 3Department of Surgical
Research, Universitatsklinikum Carl Gustav Carus, Technische
Universitat Dresden, Dresden, Germany
Colorectal cancer (CRC) is the second most common cancer in the
Western world in terms of both incidence and mortality rate. A
positive family history of CRC is observed in about 25% of the cases.
High-penetrant germline mutations in APC, MUTYH and the mis-
match repair genes MLH1, MSH2, MSH6 and PMS2 account for less
than 5% of hereditary cases whereas in the majority of these families
the genetic defect is still unknown. In order to identify novel mod-
erate- to high-risk mutations contributing to CRC predisposition we
employed genome-wide copy number profiling using high-resolution
SNP-based arrayCGH on normal tissue DNA from 32 independent
patients with microsatellite-stable CRC without polyposis. All
patients were suspected for hereditary CRC because of their young
age at diagnosis or their positive family history for CRC. We iden-
tified small (100–160 kb) copy number anomalies in five independent
families (16%), in all the cases affecting only a single gene. None of
the genes had previously been described to be involved in colorectal
cancer susceptibility. All genomic lesions were validated with mul-
tiplex ligation-dependent probe amplification (MLPA). In four cases
we were able to establish that the aberrations were inherited from one
of the parents. Two of the genomic lesions were deletions affecting a
microRNA gene, illustrating that constitutional defects in these gene
expression regulators might be common. Interestingly, at least two of
the identified genes could be linked to pathways involved in CRC
development. In an ongoing locus-specific validation screen of
independent families with suspected familial CRC (currently * 250),
we thus far found at least one of the genes to be recurrently affected,
which strongly supports its role in CRC predisposition.
A novel cause of Lynch syndrome: heritable somatic
methylation of MSH2 due to deletion of the 30 exons
of the upstream gene
M. J. L. Ligtenberg1,2, R. P. Kuiper1, T. L. Chan3,4, M. Goossens2,
K. M. Hebeda2, M. Voorendt1, D. Bodmer1, E. Hoenselaar1,
S. J. B. Hendriks-Cornelissen2, H. G. Brunner1, A. Geurts van
Kessel1, J. H. J. M. van Krieken2, S. Y. Leung3,4, N. Hoogerbrugge1
1Department of Human Genetics, 2Department of Pathology,
Radboud University Nijmegen Medical Centre, Nijmegen,
The Netherlands; 3Department of Pathology, The University of Hong
Kong; 4Department of Pathology, St. Paul’s Hospital, Hong Kong
Lynch syndrome patients are susceptible to colorectal, endometrial
and a range of other cancers due to heterozygous inactivating muta-
tions in one of the mismatch repair genes, MLH1, PMS2, MSH2 or
MSH6. During routine diagnostics for germline mismatch repair gene
mutations, multiple patients with an MSH2-deficient tumor presented
an aberrant MLPA result of a probe, which is located 16 kb upstream
of MSH2. All these patients carried an heterozygous germline dele-
tion of 4.9 kb encompassing the last exons of EPCAM (formerly
known as TACSTD1), a gene directly upstream of MSH2 encoding
the epithelial cell adhesion molecule Ep-CAM. Due to the deletion
the transcription termination signal is lost and transcription of EP-
CAM was shown to extend into MSH2.
As antisense transcription of CpG islands may lead to methyla-
tion, we tested whether the transcription of the MSH2 promoter
would lead to methylation of its CpG dinucleotides. Indeed, the
MSH2 promoter in cis with the deletion is methylated in Ep-CAM
positive, but not in Ep-CAM negative, normal tissues, thus revealing
a correlation between transcriptional read-through of the mutated
EPCAM allele and epigenetic inactivation of the corresponding
MSH2 allele. This mechanism explains the mosaic pattern of epi-
genetic inactivation of MSH2, that we observed in successive
generations (Ligtenberg et al., Nature Genetics, 41: 112–117
(2009)). Because EPCAM is expressed in the target tissues of Lynch
syndrome, subjects with a 30 end deletion of EPCAM are offered the
standard Lynch syndrome surveillance protocol. To optimize patient
care clinical data of subjects with such a deletion that leads to loss
of one functional EPCAM allele and mosaic inactivation of MSH2
are being collected.
T cell immune response against frameshift-induced
neopeptides in HNPCC
Yvette Garbe, Matthias Kloor, Lena Ehret, Susanne Eiermann,
Peter Kienle, Hanns-Peter Knaebel
Mirjam Tariverdian, Magnus von Knebel Doeberitz Institute
of Pathology, University of Heidelberg, Heidelberg, Germany
Hereditary non-polyposis colorectal cancer (HNPCC)-associated
colorectal cancers (CRCs) display high level microsatellite instability
(MSI-H) as a consequence of mismatch repair deficiency. Features of
a pronounced immune response are a hallmark of HNPCC-associated
CRCs. Previous reports suggested that MSI-induced novel tumor-
specific frameshift peptides (FSPs) underly the high immunogenicity
of MSI-H CRCs. In a recent study, we could show that FSP-specific
peripheral T cells are frequent in patients affected by HNPCC-asso-
ciated CRCs. However, the role of immune surveillance mechanisms
and the clinical significance in HNPCC still remains unknown. To
further investigate the efficacy of FSP-specific tumor-infiltrating and
peripheral T cells in HNPCC patients with MSI-H CRC, IFN-g-
ELISpot analysis, CD107a mobilization and in vitro killing assays
were performed. TiTc isolated from HNPCC-associated CRCs spe-
cifically recognized MSI-induced FSPs and showed a strong CD107a
degranulation which correlates with tumor cell lysis of HLA-matched
MSI-H, but not microsatellite stable (MSS) CRC cells.
Moreover, FSP-specific T cell responses were also present in the
majority of peripheral blood samples from patients with MSI-H, but
not MSS CRCs (p \ 0.001). FSP-specific T cell reactivity was
already detectable in the peripheral blood of healthy HNPCC family
members with MMR gene germline mutations, in contrast to healthy
control individuals without evidence for HNPCC (7/14 FSP differ-
ently recognized, p \ 0.02).
Our data strongly suggest that FSPs presented by MSI-H CRC
cells are effectively recognized by the patient’s immune system. Our
data suggest that, in vitro, FSP-specific T cells are capable of rec-
ognizing and killing MSI-H CRC cells. These results are compatible
with a prominent role of immune surveillance contributing to the
clinical course and eventually the limited penetrance of the HNPCC
syndrome. These observations are of high relevance for the devel-
opment of FSP-based vaccination approaches, particularly for the
preventive application in HNPCC mutation carriers.
Abstracts 723
123
MTHFR 677 C[T and 1298 A[C polymorphisms
and the age of onset of colorectal cancer in HNPCC
R. J. Scott, S. G. Reeves, C. Meldrum, C. Groombridge,
A. D. Spigelman, J. Suchy, G. Kurzawski, J. Lubinski, P. McElduff
University of Newcastle, Division of Genetics, Hunter Area
Pathology Service, Newcastle, Australia
HNPCC or Lynch Syndrome is associated with an increased risk of
developing an epithelial malignancy. There is considerable variability
in disease expression observed in this syndrome which is thought to
be due to a combination of genetic and environmental factors.
Alterations in the kinetics of MTHFR due to the presence of
polymorphisms in the MTHFR gene have been associated with an
increased risk of CRC. Two common single nucleotide polymor-
phisms (SNPs) located within the MTHFR gene, 677 C[T and 1298
A[C that alter the function of the encoded protein have been the
focus of many studies into CRC risk outside of the context of an
inherited predisposition to disease.
A total of 417 HNPCC patients were genotyped for the 677 C[T
and 1298 A[C SNPs to determine if there exists an association with
the age of disease onset of colorectal cancer. Associations in disease
risk were further investigated using Kaplan–Meier survival analysis
and Cox hazard regression.
The average ages of disease diagnosis were found to be different
between individuals harbouring either one of the MTHFR polymor-
phisms. Both Kaplan–Meier and Cox hazard regression analysis
revealed a more complex relationship between the two polymor-
phisms and the age of CRC onset. Kaplan–Meier survival analysis
revealed that compound heterozygotes for the two SNPs developed
CRC 10 years later than those carrying only the wild type alleles.
Hereditary nonpolyposis colorectal cancer in 688
families: mutations, age of diagnosis and cancer
incidence
Bente A. Talseth-Palmer1,2, Mary McPhillips3, Cliff Meldrum3
Claire Groombridge4, Allan D. Spigelman5 and Rodney J. Scott1,2,3
1School of Biomedical Sciences, University of Newcastle, NSW,
2308, Australia; 2Hunter Medical Research Institute, John Hunter
Hospital, Newcastle, NSW, 2305, Australia; 3Hunter Area Pathology
Service, John Hunter Hospital, Newcastle, NSW, 2305, Australia;4Hunter Family Cancer Service, Hunter New England Health, NSW
2305, Australia; 5St Vincent’s Hospital Clinical School, Sydney,
NSW 2010, Australia
Hereditary nonpolyposis colorectal cancer (HNPCC) is an autosomal
dominant inherited cancer syndrome associated with germline muta-
tions in DNA mismatch repair (MMR) genes, with the majority of
mutations affecting hMLH1 and hMSH2. The primary function of
MMR genes is to eliminate base–base mismatches and insertion–
deletion loops which arise as a consequence of DNA polymerase
slippage during DNA replication. HNPCC is characterized by early-
onset epithelial cancers, with patients reporting to have an 80%
lifetime risk of developing colorectal cancer (CRC), but are also at
risk of developing cancer in a variety of other organs that include the
endometrium, stomach, ovary, bladder, pancreas and the urinary tract.
Disease penetrance estimates have not been accurately assessed and
there is currently doubt about the true incidence of disease as well as
age dependent probabilities of cancer expression.
From 1997 to 2007, 1376 individuals (belonging to 688 families)
have been tested for HNPCC at the Hunter Area Pathology Service
(HAPS). Of these, 384 tested negative for a predictive test and
therefore do not have HNPCC and 479 tested positive for mutations in
one of the known genes (hMLH1, hMSH2, hMSH6 and PMS2). 29%
of the families submitted for mutation analysis were found to harbour
a causative change in one of the 4 genes. Missense mutations of
unknown significance were found in 12% of the families and muta-
tions were not identified in 59% of the families tested. Average age of
diagnosis of CRC is lower in mutation positive individuals (42 years
for hMLH1, 44 years for hMSH2 and 45 years for hMSH6 mutation
carriers than in mutation negative individuals 52 years).
Of the families with available information on cancer types, there
were: 71 probands with a hMLH1 mutation, 84 with a hMSH2
mutation, 17 with a hMSH6 mutation, 3 with a PMS2 mutation, 68
with a missense mutation and 352 mutation negative probands. The
incidence of extracolonic cancers was calculated to determine which
cancers were over-represented in the Australian HNPCC population.
Typically, endometrial cancer was over-represented (78 families with
causative changes in one of the MMR genes). Other cancers were also
observed at frequencies typical for HNPCC. However, our previous
data indicating an over-representation of breast cancer in 95 families
was confirmed in our updated dataset of 688 families.
This updated dataset can now be used to better define disease
penetrance and to accurately assess age specific disease incidence.
This knowledge will help in providing accurate cancer risk assess-
ments to patients harbouring causative mutations in MMR genes.
Base excision repair pathway in microsatellite stable
tumors from hereditary non-polyposis colorectal cancer
Trinidad Caldes, P. Garre1, V. Briceno1, O. Valentin1, R. Xicola2,
X. Llor2, M. de la Hoya1, T. Caldes1
Hospital Clinico San Carlos, Madrid, Spain
Base excision repair (BER) pathway is the most important cellular
protection mechanism responding to oxidative DNA damage. MYH,
OGG1 and MTH1 are members of BER, and MYH germline muta-
tions were recently identified in patients with multiple adenomas and
colorectal cancer. A subset of colorectal cancers (CRCs) arises in
families that, despite fulfilling clinical criteria for Hereditary Non-
Polyposis Colorectal Cancer (HNPCC), do not show evidence of a
mismatch repair (MMR) deficiency.
Aims To identified genetic variants in MYH, OGG1 and MTH1
genes present in HNPCC families without MMR deficiency (HNPCC-
MSS).
Methods A total of 45 DNAs from HNPCC-MSS index cases, were
screened for all coding sequences of OGG1 and MTH1 by direct
sequencing. S326C and IVS5-15C[G (OGG1) and D142D (MTH1)
polymorphisms were also typed in 353 DNAs from control individ-
uals by TaqMan assay technology. G382D and Y165D at the MYH
gen were studied previously (Peterlongo et al. 2006).
Results Three new variants were detected R46Q, A95A and G308G
in OGG1 gene and two in MTH1, the V106M and D122D that was
detected in two families. The frequencies of the SNPs, S326C and
IVS5-15C[G in OGG1 and D142D in MTH1 were 38, 35 and 56%
respectively in HNPCC-MSS index cases and 40, 35 and 36% in the
control population. The reported variant R154H in OGG1 was not
observed in Spanish population. A statistical analysis (two-sided X2
or Fisher test) was performed comparing frequencies between case
and controls. The two variants in the OGG1 gene were not signifi-
cantly different but the variant D142D in MTH1 gene was found to be
significantly different between HNPCC-MSS cases and controls (OR:
12.37, p = 0.016). The genetic variants R46Q, A95A in the OGG1
gene, segregate with colorectal cancer in the family by the contrary
724 Abstracts
123
variants D122D and V106M in MTH1 gene didn’t segregate. We
couldn’t established the segregation for the variant G308G.
Conclusions This study suggests that genetic variants in OGG1 and
MTH1 may explain by different ways the susceptibility to CRC in
HNPCC-MSS patients. Further studies are required to confirm the
reported associations.
An important role for RNA based mutation scanning
in the mismatch repair gene PMS2
H. M. van derKlift, C. M. Tops2, E. C. Bik2, M. W. Boogaard2,
H. Morreau3, F. Hes2, P. Devilee1,3, J. T. Wijnen1,2
Heterozygous mutations in the Mismatch Repair gene PMS2 are
involved in Lynch Syndrome while bi-allelic mutations are found in
Constitutional Mismatch Repair Deficiency Syndrome patients.
Mutation scanning in PMS2 is complicated by the presence of many
pseudogenes in the Human Genome, with 14 copies involving PMS2
exon 1–5 on chromosome 7q and one copy (PMS2CL) of the 30 end
(exon 9 and 11–15) about 700 kb centromeric from PMS2 on 7p.
Recently it has been recognized that frequent recombination, either
gene conversion or cross-over, between PMS2 and PMS2CL has
occurred and is still ongoing. This makes mutation detection based on
PMS2 specific nucleotides highly unreliable, especially, as we have
shown, in the region encompassing exon 12–30 UTR.
Here we propose an alternative strategy for mutation detection in
PMS2 in which RT–PCR plays an important role either as initial
scanning or as confirmation of mutations found in genomic DNA. As a
reliable source of RNA we use short term cultured lymphocytes in
which Nonsense Mediated RNA decay can be inhibited. The cDNA is
amplified in two overlapping amplicons spanning the entire PMS2
gene, thereby circumventing pseudogene sequences and gene con-
version events. With this strategy we found in 26 Lynch Syndrome
families 19 different pathogenic mutations of which 15 were detectable
with RT–PCR, the other 4 being large deletions that cross one or both
borders of the gene. Two out of 19 mutations were missed with MLPA,
exon-by-exon or Long Range PCR and could be detected with RT–
PCR and Southern Blot only: a genomic deletion of exon 14 and a large
2 kb insertion of an SVA repeat in intron 7 resulting in an mRNA with
a spliced-in fragment of 71 bp. We conclude that in our experience
RT–PCR represents an effective tool for mutation scanning in PMS2.
Early detection of upper gastrointestinal cancers
by endoscopy in patients with Lynch syndrome
Masami Arai, Toshiharu Yamaguchi, Masatoshi Oya, Junko Fujisaki,
Masahiro Igarashi, Kensuke Kuraoka, Hiroshi Takahashi, Tetsuichiro
Muto
The Cancer Institute Hospital of JFCR, Tokyo, Japan
Background Colonoscopy is considered useful for cancer surveil-
lance in patients with Lynch syndrome, but the diagnostic role of
upper gastrointestinal endoscopy remains unclear. At our hospital,
patients with Lynch syndrome are recommended to undergo annual
screening by upper gastrointestinal endoscopy and colonoscopy (plus
gynecological examination in women). We summarize the outcomes
of surveillance by upper gastrointestinal endoscopy.
Patients profiles and methods 41 patients (17 males, 24 females,
average age at entry 49.3 years) with Lynch syndrome satisfying the
Amsterdam criteria II or with confirmed pathogenic mutations
(including 1 epimutation in the promoter region of hMLH1) in either
MMR gene were observed from 2005 to 2006. Upper gastrointestinal
endoscopy was performed once every 1–2 years. Tumor incidence
and outcomes were studied. We used the incidence of tumor detection
on cancer screening by upper gastrointestinal endoscopy in our hos-
pital as a control.
Results Among 41 patients, 5 had a history of cancer (4 gastric
cancer, 1 cancer of the ampulla of Vater). A total of 107 upper gas-
trointestinal examinations were performed within 4 years. 8 lesions
(4 gastric cancers, 2 duodenal cancers, 1 SMT of the duodenum, and 1
severe dysplasia of the esophagus; incidence 7.5%) were detected. All
tumors could be detected at curative stage, and 3 were resected
endoscopically. The control tumor incidence was 0.82%.
Conclusion Regular upper gastrointestinal endoscopy can facilitate
early cancer detection in Lynch syndrome. The 2nd portion of the
duodenum should be examined to avoid missing duodenal lesions.
Adenomas of the stomach and duodenum as well as the colorectum
may be precancerous in Lynch syndrome. Because some cancers
develop rapidly, upper gastrointestinal endoscopy should be per-
formed every 1–2 years in patients with Lynch syndrome.
Primary peritoneal cancer following bilateral salpingo-
oophorectomy in two patients with Lynch syndrome
Kathleen M. Schmeler, Molly S. Daniels, Pamela T. Soliman, Russell
R. Broaddus, Michael T. Deavers, Thuy M. Vu, George J. Chang,
Karen H. Lu
MD Anderson Cancer Center, Houston, Texas, USA
Background Women with Lynch syndrome have a 40–60% lifetime
risk of endometrial cancer and a 7–12% lifetime risk of ovarian
cancer. Risk-reducing surgery including hysterectomy and bilateral
salpingo-oophorectomy is currently recommended once childbearing
is complete. Unlike women with BRCA mutations, there have been
no reported cases of primary peritoneal cancer following bilateral
salpingo-oophorectomy (BSO) in women with Lynch syndrome. The
objective of this study was to describe two cases of primary peritoneal
cancer following BSO in women with Lynch syndrome.
Cases The first patient was a 44 year-old woman who underwent
hysterectomy with BSO for benign disease. She presented 12 years later
with a pelvic mass and was diagnosed with a high-grade serous primary
peritoneal cancer. Genetic testing showed a mutation in the MSH2 DNA
mismatch repair gene. The second case was 58 year-old woman who
had a hysterectomy and BSO for endometrial cancer. She developed a
high-grade serous primary peritoneal cancer 8 years later and was
found to have a mutation in the PMS2 DNA mismatch repair gene.
Conclusion Women with Lynch syndrome should be counseled that
they are at risk for developing primary peritoneal cancer despite
undergoing gynecologic cancer risk-reducing surgery. The magnitude
of this risk remains to be determined.
Clinical and genetic characteristics of Japanese
HNPCC
Yoichi Furukawa, Teruhiko Yoshida, Yusuke Nakamura, Yoshihiro
Moriya
Institute of Medical Science, The University of Tokyo, Tokyo, Japan
To uncover the clinical and genetic characteristics of Japanese
HNPCC, we performed a collaborative study of HNPCC registry and
Abstracts 725
123
genetic testing project in Japan. Patients who fulfilled with modified
Amsterdam criteria that included gastric cancer as an HNPCC-related
tumor were enrolled in this study.
A total of 101 patients were subjected to genetic testing after the
informed consent was obtained. We analyzed genetic alterations in
MSH2, MLH1, and MSH6, three major responsible genes by PCR
and direct sequencing and MLPA. As a result, we identified 56
pathogenic mutations in the 101 cases. Among the 56 alterations,
31, 23, and 2 mutations were found in MLH1, MSH2, and MSH6,
respectively. Although 47 mutations were detected in 70 patients
who fulfilled with revised Amsterdam criteria, the remaining 9
patients were found in 31 subjects who might not be enrolled if
gastric cancer was not taken as a related tumor. This result suggest
that inclusion of gastric cancer increases the sensitivity of diagnosis
but decreases specificity, leaving further studies for the development
of systematic screening system.
Expectedly, we observed that early onset and multiple HNPCC-
related tumors were associated with mutation. In addition, we found
six subjects who harbored the same deletion of MLH1 exon 5.
Further information on their family trees has uncovered that two of
the five probands were relatives in a large pedigree. These data may
be useful for the development of screening system to Japanese
HNPCC.
Identification of the first Spanish founder mutations
in the MLH1 gene
M. Pineda1,*, E. Borras1,*, I. Blanco2, G. Llort3, T. Caldes4,
M. Urioste5, C. Martınez-Bouzas6, B. Grana7, J. Balmana8,
A. Torres9, T. Ramon y Cajal10, J. Sanz9,11, M. Duran11,
S. Castellvı-Bel12, S. Gonzalez1, C. Lazaro1, G. Capella1
1Laboratori de Recerca Translacional, Institut Catala d’Oncologia,
IDIBELL, Hospitalet de Llobregat, Spain; 2Unitat de Consell Genetic,
Institut Catala d’Oncologia, IDIBELL, Hospital Duran i Reynals,
Hospitalet de Llobregat, Spain; 3Unitat de Consell Genetic, Institut
Catala d’Oncologia, Hospital Germans Trias i Pujol, Badalona, Spain;4Laboratorio de Oncologıa Molecular, Hospital Clınico Sant Carlos,
Madrid, Spain; 5Departamento de Genetica Humana, Centro Nacional
de Investigaciones Oncologicas, Madrid, Spain; 6Laboratorio de
Genetica Molecular, Hospital de Cruces, Bizkaia, Spain; 7Unitat
d’Avaluacio del Risc de Cancer i Consell Genetic, Institut Catala
d’Oncologia, Hospital Universitari Josep Trueta, Girona, Spain;8Servei d’Oncologia Medica, Hospital Vall d Hebron, Barcelona,
Spain; 9Unitat de Consell Genetic, Hospital Universitari Sant Joan,
Reus, Spain; 10Servei d’Oncologia Medica, Hospital de la Santa Creu
i Sant Pau, Barcelona, Spain; 11Institut d’Oncologia Corachan-Centre
Medic, Barcelona, Spain; 12Servei de Gastroenterologia, Institut de
Malalties Digestives i Metaboliques, Hospital Clınic, Barcelona,
Spain; 13Instituto de Biologıa y Genetica Molecular, Valladolid,
Spain
Lynch syndrome is an autosomal disorder caused by germline
mutations in any of four DNA mismatch repair genes: MLH1, MSH2
MSH6 or PMS2. Clinically it is characterized by the development of
early-onset colorectal cancers and an increased risk of other cancers.
Lynch syndrome tumors are associated with microsatellite instability
and loss of mismatch repair proteins expression. In some populations,
founder mutations appear to explain a substantial fraction of Lynch
syndrome cases.
We report here the identification of two frequently occurring
germline substitutions of the MLH1 gene in Spanish families:
c.306+5G[A and C.1865T[A (p.Leu622His). The c.306+5G[A
mutation was identified in 17 unrelated families from the north-east
of Spain, whereas the C.1865T[A mutation was found in 12 unre-
lated families coming from the region of Jaen, at the south of Spain.
Both mutations segregate with the disease, and tumors show
microsatellite instability and loss of expression of MLH1 protein.
We demonstrate that c.306+5G[A is a pathogenic mutation affect-
ing the mRNA processing. Experimental approaches are being
performed to elucidate the pathogenicity of the C.1865T[A
(p.Leu622His) mutation.
To determine possible founder effects on the identified Spanish
MLH1 mutations, haplotype analysis was performed using three
MLH1 SNPs and seven microsatellite markers, spanning 12 Mb on
chromosome 3. The haplotype analysis demonstrated two common
haplotypes associated to c.306+5G[A and C.1865T[A MLH1 muta-
tions in the families. The estimation of the age of the c.306+5G[A and
C.1865T[A mutations was of 36–70 and 5–65 generations ago,
respectively. Mutations c.306+5G[A and C.1865T[A (p.Leu622His)
of MLH1 gene are the first founder MLH1 mutations identified in
Spain. This finding should be taken into consideration when designing
molecular diagnostic strategies in the Spanish Lynch syndrome
families.
Assessing BRAF V600E usefulness in the analytical
algorithm of Lynch syndrome
M. Gausachs1, M. Pineda1, M. Menendez1, C. Lazaro1, I. Blanco2,
S. Gonzalez1, G. Capella1
1Laboratori de Recerca Translacional, 2Unitat de Consell Genetic,
Institut Catala d’Oncologia, IDIBELL, l’Hospitalet de Llobregat,
Spain
BRAF is a kinase-encoding gene from the RAS/RAF/MAPK path-
way. The hotspot BRAF V600E is present in about 10–15% of
sporadic colorectal cancers (CRC) and strongly associates with
microsatellite instability (MSI). In MSI tumors with loss of MLH1
expression BRAF mutation detection is used in the selection of
candidates to have Lynch syndrome, since V600E is believed to be
diagnostic of sporadic tumors except for PMS2 families.
The aim of this study was to assess the usefulness of BRAF V600E
in the analytical algorithm of tumors from candidates to mismatch
repair (MMR) germline screening.
A set of 157 tumors from individuals with family history of CRC
attended at our Cancer Genetic Counseling Unit [109 MSI and 48
MSS (microsatellite stable)] were included. SNuPE test was opti-
mized to detect BRAF V600E in DNA from paraffin-embedded
tumors with an analytical sensitivity of 1:200. Assay was initially
validated in a set of selected sporadic CRC [24 MSI and 49 MSS]. In
these tumors BRAF mutation was identified in 5 of 24 (20%) MSI and
2 of 49 (4%) MSS tumors.
BRAF mutation was assayed in 15 of 157 (9.5%) familial tumors
[12 of 109 (11%) MSI; 3 of 48 (6%) MSS]. Restricting the analysis
to MSI tumors, MMR germline mutations were found in 63 (43
MLH1 and 18 MSH2 and 2 MSH2) of the 109 MSI CRC. One
BRAF V600E mutation was detected in a tumor from a germline
MLH1 carrier and the remaining 11 mutations were found in the
cases without detectable germline mutation. BRAF sensitivity was
assessed in 51 cases with loss of MLH1 protein expression. All
BRAF mutations associated with loss of MLH1 expression, showing
a sensitivity of 39% and a specificity of 96% for depiction of
sporadic tumors.
This is the first report of the coexistence of somatic BRAF V600E
and MLH1 germline mutation. Our findings question its role in the
diagnostic algorithm of Lynch syndrome.
726 Abstracts
123
Diagnostic microsatellite instability testing in Lynch
syndrome: different tumours, different markers
Kenneth Porter, Nathan Curtis, Lisa Happerfield, Mark J. Arends,
Ian M. Frayling
University of Southampton School of Medicine, Southampton
General Hospital, Southampton, UK
Microsatellite instability (MSI) testing plays a key role in the
molecular diagnosis of Lynch Syndrome (LS), together with mis-
match repair protein expression by immunohistochemistry, and
germline mutation detection. Because different microsatellite markers
have different sensitivities to detect MSI in different tumours, we
have examined marker sensitivities in a diagnostic clinical setting.
MSI results from 285 tumours, from affected members of families
attending two cancer genetics clinics, who either met the Amsterdam
criteria or had a strong clinical suspicion of LS, were audited. A panel
of 10 microsatellites was used: 3 mono- and 7 dinucleotide repeats:
BAT25, BAT26, BAT40, D5S346, ACTC, D17S250, D13S153,
D5S406, D5S107 & D2S123. A diagnosis of LS-associated MSI was
made if[29% of markers were unstable, and 5 or more markers gave
informative results.
LS was subsequently diagnosed in 55/285 families, and the indi-
vidual and combined sensitivities of the markers (mononucleotides
alone; dinucleotides alone; all combined) in the LS and non-LS
groups were thus determined, in colorectal and non-colorectal
tumours. Because most (76%) of the tumours were colorectal in
origin, the size of the non-colorectal group did not justify sub-division
into endometrial and other non-colorectal tumours.
In those LS-associated colorectal cancers in which all three
mononucleotide markers worked, MSI could always have been
diagnosed on the basis of the mononucleotides alone, i.e. the dinu-
cleotides did not add extra information. However, in non-colorectal
tumours the mononucleotide markers were not, in themselves, always
sufficient to make a diagnosis of MSI.
We conclude that when testing colorectal tumours the marker set
could be rationalized to 5 mononucleotides, reducing costs without
compromising diagnostic efficiency. Whereas, when testing non-
colorectal tumours a set of both mono- and dinucleotide repeats
remains necessary. Further work is necessary to determine the opti-
mum marker sets to diagnose LS-associated MSI in the various types
of LS-associated tumours.
Methylation specific multiplex ligation-dependent probe
amplification in the analysis of MLH1 promoter
hypermethylation
Lars Henrik Jensen, Jan Lindebjerg, Lene Byriel, Steen Kølvraa,
Dorthe Cruger
Danish Colorectal Cancer Group South, University of Southern
Denmark and Vejle Hospital, Vejle, Denmark
Background Promoter hypermethylation can lead to transcriptional
silencing of certain genes. MLH1 is one of the main genes in the post-
replicative mismatch repair, and promoter hypermethylation of this
gene is found in sporadic mismatch deficient colorectal cancer. Lynch
Syndrome is caused by germ-line mutations in for example MLH1.
Thus, promoter hypermethylation of MLH1 can separate sporadic
mismatch deficient tumours from possible hereditary cases. The aim
of this study was to evaluate methylation specific multiplex ligation-
dependent probe amplification (MS-MLPA) in the analysis of MLH1
promoter hypermethylation and to determine the importance of the
different regions of the promoter.
Methods Two-hundred-thirty-one patients with primary colorectal
cancer were included in the study. Mismatch repair deficiency has
previously been determined by microsatellite analysis and immuno-
histochemistry. A commercially available kit was used for MS-MLPA
(ME011, MRC-Holland, Amsterdam, The Netherlands). Methylation
was determined in four different regions of the MLH1 promoter
(A through D) and in intron 1.
Results Ninety tumours (40.0%) were in some degree methylated in
one or more of the regions A–D and intron 1. All five regions were
methylated in 32 tumours (13.9%), only A in 28 (12.1%), only D in
22 (9.5%), A + B in 3 (1.3%), A + B + C in 2 (0.9%), A + D in 2
(0.9%), and A + C + D in 1 (0.4%). In all 32 tumours with methyl-
ation in five regions, immunohistochemistry was negative for MLH1
and they had a high degree of microsatellite instability. Isolated
methylation of region A, region D, or the combinations of A, B, C,
and D was not associated with microsatellite instability or loss of
MLH1 protein. Methylation in a region of intron 1 was strongest
correlated with loss of protein expression and microsatellite
instability.
Conclusion MS-MLPA is an excellent tool for analysis of MLH1
promoter hypermethylation. The single most important region for
methylation analysis may be in intron 1.
Recommendation for the current endoscopic technique
in the surveillance of HNPCC gene carriers
R. Huneburg1, T. Sauerbruch1, C. Lamberti1
1Department of Internal Medicine I
Introduction Due to the high risk of colorectal cancer in carriers of
hereditary nonpolyposis colorectal cancer (HNPCC), and due to the
shortened adenoma–carcinoma sequence, good colonoscopic surveil-
lance is obligatory in these patients. Up to 25% of small and flat
adenomas are missed by standard colonoscopy (SC). Therefore an
endoscopic approach is needed which is able to detect small adenomas.
Methods All endoscopic studies that met the following inclusion
criteria:
1. Screening of HNPCC patients
2. Comparison of different endoscopic techniques.
Results To date, there are 4 studies available, 3 published, 1 in
progress. In two studies, SC was compared to chromocolonoscopy
(CC), in one SC was compared to narrow-band-imaging colonoscopy
(NBI), in the last one, a two-armed study, SC was compared to CC as
well as CC to NBI.
A total of 105 patients were screened by comparing SC to CC,
62 patients by comparing SC to NBI and 62 patients by comparing
CC to NBI. In all CC studies indigo carmine dye was used. The
percentage of HNPCC gene mutation carriers differed a lot (84/50%/
13/90%) as well as the rate of patients who had a colonoscopy been
performed before inclusion into the study (32/56%/100/92%). Three
studies showed a significant increase of adenoma detection while
using CC (SC 25 adenomas/CC 56). One study showed a significant
increase while using NBI (SC 25/NBI 21). One study revealed that
CC detected significantly more adenomas than NBI (NBI 10/CC
38). In all four studies no randomized controlled trial was
performed.
Conclusion CC is superior to standard and NBI colonoscopy in
detection of polypoid lesions in patients with HNPCC. Further studies
are needed, preferably randomized controlled trials.
Abstracts 727
123
Colorectal cancers show distinct mutational spectra
in members of the WNT pathway according
to anatomical localization and geneticinstability
Cristina Albuquerque5, Celia Baltazar1, Bruno Filipe1, Filipa Penha1,
Teresa Pereira2, Ron Smits3, Marılia Cravo4, Pedro Lage4, Paulo
Fidalgo4, Isabel Veiga5, Jose Silva Ramos6, Isabel Fonseca2,
Carlos Nobre Leitao4, Riccardo Fodde3
1Centro de Investigacao de Patobiologia Molecular (CIPM), 2Servico
de Anatomia Patologica e, 4Servico de Gastrenterologia, Instituto
Portugues de Oncologia de Lisboa Francisco Gentil, EPE, Lisboa,
Portugal; 3Department of Pathology, Josephine Nefkens Institute,
Erasmus MC, Rotterdam, The Netherlands; 5Servico de Genetica,
Instituto Portugues de Oncologia do Porto Francisco Gentil, EPE,
Porto, Portugal; 6Servico de Gastrenterologia, Hospital dos Capuchos,
Lisboa, Portugal
Constitutive activation of the canonical WNT signal transduction
pathway plays a rate-limiting role in colorectal cancer onset and
progression. However, it is yet unclear whether the mutation spectra
in WNT genes vary depending on the sporadic or hereditary nature of
the tumors, microsatellite instability (MSI) status, and localization
along the gastro-intestinal tract. To clarify this, we have analyzed the
APC (adenomatous polyposis coli), CTNNB1 (beta-catenin) and
AXIN2 (conductin) genes for somatic mutations in 52 colorectal
tumors and performed a meta-analysis of previous studies. In our
cohort, mutations were detected in the majority of sporadic micro-
satellite stable (MSS; 80%) and MSI-High (MSI-H; 60%) carcinomas
and HNPCC (hereditary non-polyposis colorectal cancer) colorectal
tumors (74%). Notably, significant differences were found in muta-
tion spectra among the different groups. Truncating APC mutations
encompassing at least two beta-catenin down regulating motifs (20
a.a. repeats) were significantly more frequent among sporadic MSI-H
and HNPCC cases than in MSS tumors (p = 0.027). In the latter,
APC mutations encompassing only one or none of the 20 a.a. repeats
were prevalent. Also, beta-catenin mutations were almost exclusively
detected in MSI-H tumors, being more frequent in HNPCC than in
sporadic lesions (40% vs. 10%). With respect to anatomical locali-
zation, APC mutations encompassing at least two 20 a.a. repeats were
also significantly more frequent in proximal tumors than in tumors
located in the descending/sigmoid colon (p = 0.047). The meta-
analysis confirmed our results, leading to more significant data. In
conclusion, the observed spectra of WNT gene mutations in HNPCC
and sporadic MSI-H tumors are likely to result from selection of
specific levels of beta-catenin signaling, optimal for tumor formation
in specific anatomical locations and genetic backgrounds and may
contribute for distinct clinical features and prognosis.
Familial colorectal cancer type X tumours present
frequently chromosomal instability, APC allelic loss
and specific KRAS mutations
Ines Francisco1, C. Albuquerque1, H. Belo1, B. Filipe1, I. Vitoriano1,
J. Dinis1, P. Lage2, I. Claro2, S. Ferreira2, P. Rodrigues2, C. Nobre
Leitao2
1Centro de Investigacao de Patobiologia Molecular (CIPM), 2Servico
de Gastroenterologia, Instituto Portugues de Oncologia de Lisboa
Francisco Gentil, EPE (IPOLFG, EPE), Portugal
In a fraction of families fulfilling the Amsterdam Criteria (AC) for
hereditary non-polyposis colorectal cancer, DNA mismatch repair
gene mutations are not found and colorectal cancers (CRC) are mostly
localized in the distal colon and frequently do not present microsat-
ellite instability (MSI). These families have been designated as
familial colorectal cancer type X (FCCX). We aimed at characterize a
group of FCCX families for both clinical and molecular features.
Twelve patients with CRC and/or adenomas, from 10 FCCX families
were included and 8 CRC and 4 adenomas were analyzed. Chromo-
somal instability (CIN) was evaluated in the 8 CRC by the analysis of
loss of heterozygosity (LOH) at 5q, 17p and 18q. APC (point muta-
tion/LOH) and KRAS mutations were analyzed in the 12 tumours.
When compared with Lynch syndrome, FCCX families presented a
lower frequency of CRC and extra-colonic tumours from the Lynch
spectrum but showed a higher frequency of adenomas. Among CRC,
5/8 were classified as presenting CIN. LOH of APC was detected in
7/11 tumours, including all CIN carcinomas. APC and KRAS muta-
tions were identified in 4/11 and 6/12 tumours, respectively, mostly in
tumours presenting CIN or LOH of APC. Considering the high fre-
quency of LOH of APC and the higher prevalence of adenomas in
FCCX, we searched for APC germline mutations and found the
E1317Q mutation in one family. In conclusion, FCCX families
present specific clinical and molecular features and may be divided in
two distinct groups: one that present CIN tumours, with frequent LOH
of APC and a specific KRAS mutation signature and a second group
where tumours do not present CIN and where these alterations may be
less frequent. The identification of the E1317Q mutation in one of the
families from the former group suggests that some APC missense
mutations may contribute for these cases.
Analysis of colonic and extra-colonic tumors in Chilean
patients with Lynch syndrome
T. Claudio Heine1, M. De la Fuente1, K. Alvarez1, E. Pinto1,
U. Kronberg1, A. M. Wielandt1, P. Orellana1,2, P. Carvallo2,
F. Lopez-Kostner1
1Colorectal Surgery Unit, Oncology and Molecular Genetics
Laboratory, Clınica las Condes, Santiago, Chile; 2Human Molecular
Genetics Laboratory, Pontificia Universidad Catolica de Chile,
Santiago, Chile
Background The Lynch Syndrome or Hereditary Non Polyposis
Colorectal Cancer (HNPCC) represents up to 5% of all cases of
colorectal cancer. Theses patients have extracolonic tumors also in
various organs such as endometrial, small intestine, ureter and renal
pelvis. All this tumors have been included in different clinical criteria
for patient selection and diagnosis.
Objective To describe and characterize colonic and extra-colonic
tumors in Chilean families with Lynch syndrome.
Materials and methods From hereditary intestinal tumor registry, we
selected families that meet the clinical criteria of Amsterdam or
Bethesda (with microsatellite instability). In all families, we evaluated
the presence of colorectal and extracolonic tumors.
Results We identified 14 families (83 patients) who meet the
Amsterdam criteria and 7 families (23 patients) who meet the Bethesda
criteria (with microsatellital instability). A total of 143 tumors were
identified, 82 colorectal (57%) and 64 extracolonic (43%). The dis-
tribution of colorectal tumors showed a higher frequency of right colon
(54%) followed by left colon (22%) and rectum (24%). Regarding
extracolonic tumors, the most frequent was endometrial cancer (13%)
followed by gastric cancer (12%) and breast cancer (10%). We also
note that other cancers appeared before colorectal cancer, like endo-
metrial cancer (75%) and breast cancer (67%). Twenty-eight patients
had more than one tumor (26%) and 7 presents more than 3 tumors
(7%) with a higher proportion in females (71%).
728 Abstracts
123
Conclusions The results confirm the high incidence of colonic and
extracolonic tumors in Chilean families with Lynch Syndrome. It also
highlights the high incidence of gastric tumors, which are not con-
sidered within the Amsterdam criteria II, and breast tumors, which are
not included within the Bethesda criteria.
Mutation analysis of the MMR genes in patients
with suspicion of HNPCC from Castilla-Leon (Spain:
Identification of a novel and recurrent deletion
in MSH2)
L. Perez-Cabornero, E. Velasco, M. Infante, D. J. Sanz, A. Acedo,
L. Hernandez, N. Martinez, E. Lastra, C. miner, M. Duran
Instituto de Biologıa y Genetica Molecular (IBGM), Valladolid, Spain
Introduction Hereditary non-polyposis colorectal cancer (HNPCC)
is associated with germline mutations in DNA mismatch repair genes
(MMR genes), predominantly MLH1, MSH2 and MSH6. Defects in
the MSH2 gene may account for about 40% of HNPCC cases. The
aim of the study is to describe the mutational spectrum in 140 non-
related families with suspicion of HNPCC from Catilla-Leon.
Methods Point mutations in the MLH1, MSH2 and MSH6 genes were
screened by heteroduplex analysis by capillary array electrophoresis
(HA-CAE) followed by direct sequencing. Genomic rearrangements
were assessed by multiplex ligation-dependent probe amplification
(MLPA) and the positive results were confirmed by RT–PCR.
Results Fourty-three different MMR gene alterations were detected.
Among these, 9 mutations in 13 families are considered pathogenic and
an additional 9 variants from 16 families are considered to represent
variants of unknown pathogenicity. About 40% (7/18) of the pathogenic
and UV mutations were novel: one complex frameshift mutation (MLH1
c.2284-2287delACCT; 2284-2313ins30), one splicing mutation (MSH2
C.1661G[A), three unknown missense variants (MLH1 C.1574G[A;
MSH6: c.4003 A[C, c.3425C[T and c.100g[C) and a large genomic
deletion in MSH2 removing exon 4–8. The different MMR genes con-
tribute to 61% (MSH2), 31% (MLH1), and 8% (MSH6) of the mutations.
About 50% of pathogenic mutations in MSH2 are large genomic dele-
tions. Most of them were present in a single family. Interestingly, we
found a novel and recurrent deletion exon 4–8 in MSH2, previously
recognize, was causative in 31% (4/13) of the MMR mutant families.
Conclusion These data indicate that: (1) the majority of germline
mutations in HNPCC families in Castilla-Leon affect the MSH2
locus; (2) In our population we have identified a high proportion of
novel mutation; (3) The identification of a new recurrent deletion is
relevant to the molecular diagnosis of HNPCC in this region of Spain.
In silico analyses and experimental validation of MLH1
and MSH2 splice site and missense mutations
Beate Betz1, Stephan Theiss2, Murat Aktas1,3, Carolin Konermann4,
Timm O. Goecke1, Gabriela Moslein5, Heiner Schaal4, Brigitte
Royer-Pokora1
1Institut fuer Humangenetik, Universitaetsklinikum Duesseldorf,2RESULT GmbH, Duesseldorf, 3Institut fuer Transplantations-
diagnostik und Zelltherapeutika, Universitaetsklinikum Duesseldorf,4Institut fuer Virologie, Universitaetsklinikum Duesseldorf5Klinik fuer Chirurgie, St Josefs-Hospital Bochum-Linden
Hereditary non-polyposis colorectal cancer is caused by inactivating
mutations of DNA mismatch repair genes, mainly MLH1 and MSH2.
Identification and characterization of those mutations is essential for
genetic counseling. However the clinical effect of missense and splice
site mutations remains unclear, until the pathogenic effect can be
demonstrated. Missense mutations, lying in cis-acting sequences of
splicing regulatory proteins (splicing enhancers/silencers) can cause
aberrant splicing.
We assessed ten splice site mutations and nine missense mutations
outside splice sites in MLH1 and MSH2 and experimentally analyzed
their effect on aberrant splicing at the RNA level. We additionally
tested and compared the reliability of several web-based programs for
the prediction of splicing outcome for these mutations. We used four
well-established algorithms for computational scoring of 50 and/or 30
splice sites (Shapiro & Senapathy, MaxEnt score, HBond score,
SD-score). For the prediction of putative exonic splicing enhancers/
silencers, we tested five common web-based resources representing
different algorithms (ESE-Finder 3.0, Rescue-ESE, FAS-ESS, PESX,
ESR-Search).
Our study revealed current limits of the applied computational
tools in the diagnostics of possibly pathogenic splicing mutations.
Seven splice site mutations caused aberrant splicing: in six cases
exon skipping and in one case activation of a cryptic splice site was
observed. One 50ss mutation caused the creation of a new overlapping
50ss. Splice site strength assessments showed a high consistency both
among the four different algorithms examined and compared to the
RNA-based data. However, some programs exhibited exemplary
weakness in specific positions and the type of aberrant splicing (exon
skipping or activation of a cryptic splice site) can not yet be predicted
by any of the examined algorithms.
None of the experimentally tested missense mutations in the
MLH1 and MSH2 genes actually influenced splicing in our RNA-
assay. In contrast, we found considerable divergence in the software-
predicted splicing regulatory elements.
Integrated evaluation of unclassified variants
in the mismatch repair genes
Pastrello Chiara, Pin Elisa, Agostini Marco, Barana Daniela,
Fornasarig Mara, Tibiletti Maria Grazia, Quaia Michele, Ponz de
Leon Maurizio, Pucciarelli Salvatore
Viel Alessandra Centro di Riferimento Oncologico, Aviano (PN),
Italy
Genetic testing of mismatch repair genes for the diagnosis of Lynch
Syndrome is routinely carried out in our laboratory. The result of a
14 year-long screening revealed, beside the most informative non-
sense, frameshift and large-deletion mutations, also several missense,
silent and deep intronic variants with uncertain pathogenic meaning
(unclassified variants). To clarify the role of these variants, we per-
formed an integrated analysis evaluating:
Correlation to clinical data
Cosegregation with disease
Presence of tumour molecular markers (MSI, IHC, LOH)
Predictive value of multiple in silico analyses
cDNA splicing alterations
cDNA allelic imbalance compared to gDNA
Presence of the variant in healthy controls
An arbitrary ‘‘pathogenicity value’’ was assigned to every parameter
and then summed to obtain a final score for each variant. These sums
were subdivided in 3 value ranges: probably pathogenic, probably non
pathogenic and intermediate.We analyzed 42 variants: 15 MLH1, 12
MSH2, 5 MSH6 and 10 PMS2; 26 were missense, 8 were silent and 8
were deep intronic.
Abstracts 729
123
p.Pro349Arg, p.Met688Arg and p.Cys697Arg for MSH2, p.Gly-
67Arg, p.Thr82Ala, p.Arg265His and p.Lys618Ala for MLH1 and
p.Ser46Ile for PMS2 resulted probably pathogenic. On the contrary, we
had evidence of neutrality for a large number of variants, among which
the common p.Gly322Asp and c.1077-10 T[C for MSH2, and
p.Ser406Asn for MLH1 that resulted probably non pathogenic.
This simple integrated analysis of the unclassified variants helped
to clarify their pathogenic role, supporting a better estimation of the
risk and more appropriate carrier surveillance.
Practical usefulness and problem
of immunohistochemistry and microsatellite instability
test for recruitment of hereditary nonpolyposis
colorectal carcinoma patients
Noriko Motoi, Yurika Mitsuhashi, Tomoyo Kakita, Mayumi Ogawa,
Yuri Sato, Masami Arai, Yo Kato
Cancer Institute, Japanese Foundation for Cancer Research, Tokyo,
Japan
Background It is important for diagnosis of HNPCC to recruit
candidates at general hospital. Immunohistochemistry (IHC) and
microsatellite instability (MSI) test are relatively common technique,
so it might be good means of HNPCC-screening. We report our
screening system and the summary of data.
Materials and methods Since 2005, 625 specimens were examined
by IHC and MSI. IHC includes three major DNA mismatch repair
(MMR) system (MLH1, MSH2 and MSH6). MSI test includes at least
five of seven major loci (NCI panel, TGF beta-RII and BAT40).
Germline mutation of MMR genes were examined among the IHC/
MSI screened HNPCC-candidates who accepted informed consents.
Results Any loss of MMR expression by IHC was revealed in 54
samples (8.6%). 18 showed vague results whether normal or lost. Loss
of MLH1 is the most (n = 30), followed by MSH2 (19) and MSH6 (3).
Eight samples showed double loss of MSH2 and MSH6. MSI were done
in 604 cases. Three samples were omitted due to technical problems. 18
samples were examined by IHC alone. MSI was high in 55 samples
(9.1%), vague in 9 (1.5%) and low in 540. The results between IHC and
MSI showed good correlation (p \ 0.001). The incidence of germline
mutation was 79% of IHC-lost/MSI-high cases (26/33). The most
common mutation was found in MSH2 (n = 16), followed by MLH1
(7) and MSH6 (3). Discordant between IHC and genetic test were found
in three cases. No mutation was detected in 7 IHC-lost/MSI-high cases
(21%), especially in IHC-MLH1-lost cases.
Discussion This data might be the largest series of HNPCC
screening of single institution in Japan. Sensitivity of IHC/MSI was
very high, although some cases showed non-specific IHC-reactivity.
The latter might be affected by epigenetic status or other genetic
abnormalities. Further improvement of screening methods to exclude
such non-specificity would be expected.
A homozygote splice site PMS2 mutation as cause
of TURCOT syndrome gives rise to two different
abnormal transcripts
Wenche Sjursen, Inga Bjørnevoll, Helge Myrvold,
Lars F. Engebretsen, Tore Halvorsen
Department of Pathology and Medical Genetics, St Olavs University
Hospital, Trondheim, Norway
Turcot syndrome is a rare, inherited disease predisposing to tumours
in the central nerve system and in the colorectal system.
We here describe a family in which two siblings have Turcot
syndrome, and where we have identified the underlying molecular
genotype. The proband is still alive at the age of 43. The patient was
operated at the age of 10 by brain tumour and at the age of 16 by
colorectal cancer. She has since been treated for multiple cancers
(gastrointestinal, endometrial, basal cell carcinomas), and removal of
adenomatous polyps at several occasions.
The aim of this work was to investigate if there was any specific
genotype that explains the remarkable clinical history. DNA and
RNA mutation analysis were performed for genes involved in pol-
yposis and mismatch repair. cDNA analysis for the mismatch repair
gene PMS2 showed that the patients genotype was a homozygous
splice site mutation, c.989-1 G\T, which resulted in two abnormal
transcripts, not one as expected. T he patient’s long time survival
may in part be explained by meticulous follow up by health care
professionals. The other importing factor is probably the nature of
the genotype. cDNA analysis showed that the homozygous mutation
led to two abnormal transcripts, of which one is perhaps less
detrimental.
We have detected the same mutation in heterozygote form in three
other families. It has been a discussion whether a heterozygote PMS2
mutation may be the cause of HNPCC/Lynch syndrome. We will
present these families and discuss the effect of having the variant in
heterozygous form.
Germline allele-specific expression of TGFBR1:
promoter methylation and frequency in African
Americans and in colorectal cancer with mismatch
repair deficiency
Laura Valle1,2, Tarsicio Serena-Acedo1, Sandya Liyanarachchi1,
Heather Hampel1, Ilene R. Comeras1, Stephan M. Tanner1, Albert
de la Chapelle1
1Human Cancer Genetics Program, Comprehensive Cancer Center,
The Ohio State University, Columbus, OH; 2Present Affiliation:
Laboratori de Recerca Translacional, Institut Catala d’Oncologia,
Barcelona, Spain
Most of the genetic predisposition to colorectal cancer (CRC) has not
yet been explained: While the proportion of all CRC that is associated
with genetic predisposition is expected to be as high as 35% only
around 5% has been explained molecularly so far. Up to now the
classical methods used to search for susceptibility genes in families
with high CRC aggregation (linkage analyses), have failed to identify
the genetic cause of the excess of heritability in CRC, and case–
control or cohort genome-wide association studies (GWAS) have
started to identify variants associated with very low risks and usually
located in genomic regions ‘‘empty’’ of genes.
Recently, we showed that germline allele-specific expression
(ASE) of the TGFBR1 gene occurs in 10–20% of all CRC patients
and only in 1–3% of healthy controls in a mostly Caucasian popu-
lation. If the preliminary findings are confirmed in larger studies, this
places ASE of TGFBR1 among the high-risk biological markers of
CRC predisposition with an odds ratio of at least 8.7. It is vital to
identify the cause of this partial allelic expression reduction that,
despite the efforts invested, has not been identified yet. Two major
TGFBR1 haplotypes are predominant among ASE cases, suggesting
that the genomic change is in cis, but causative germline changes
have not been identified. Moreover, the haplotype findings suggest
that the frequency of ASE might show differences between
populations.
730 Abstracts
123
The partial reduction of expression of one allele might well be
explained by germline allele-specific methylation. Methylation of
CpG sites in the promoters of genes can lead to loss of transcription
and has been repeatedly implicated in the silencing of tumor sup-
pressor genes. This hypermethylation is usually somatic, but if it
plays a role in ASE of TGFBR1, it must be heritable and occur in the
germline. Germline methylation of promoter CpG sites has been
documented (e.g. in MLH1), but it is a very rare phenomenon and its
heritability has not been convincingly proved.
Here we study germline methylation status of the TGFBR1 CpG
island in 22 ASE and 15 non-ASE individuals, not finding significant
differences. Moreover we determined the frequency of TGFBR1 ASE
and the associated haplotypes in African American (AA) CRC patients
(n = 84) and controls (n = 168), and assessed its involvement in the
predisposition to microsatellite unstable (MSI) CRC (n = 116).
To sum up, ASE of TGFBR1 is not caused by allele-specific
promoter hypermethylation, predisposes to CRC in AAs to approxi-
mately the same degree as in Caucasians, and it is not involved in the
susceptibility to CRC associated with an inherited or acquired defi-
cient mismatch repair. In AAs, the presence at lower frequency of two
Caucasian ASE-associated haplotypes suggests admixture, and the
existence of different genetic variants responsible for ASE.
Functional studies of unclassified variants of the human
mismatch repair protein MLH1 in the yeast
Saccharomyces cerevisiae
Karin Hardt1, Sven Boris Heik2, Johannes H. Hegemann2,
Brigitte Royer-Pokora1, The German HNPCC Consortium
1Institut fuer Humangenetik, Heinrich Heine Universitaet,
Duesseldorf; 2Institut fuer Funktionelle Genomforschung,
Heinrich Heine Universitaet, Duesseldorf
Genome stability is ensured by DNA-repair mechanism and is con-
served from bacteria to men. HNPCC is caused by germ line
mutations in the mismatch repair genes MLH1, MSH2, MSH6 and
PMS2. 30% of all mutations in the mismatch repair gene hMLH1 are
missense mutations. Functional analyses of proteins with amino acid
substitutions are important for the classification of missense mutations
into pathogenic or non-pathogenic variants. Due to the high homology
of the Saccharomyces cerevisiae repair system this organism can be
used as model for functional analyses of the human mismatch repair
protein hMLH1. 8 missense mutations were inserted into the ATPase
domain and 18 into the interaction domain of hMLH1. The interaction
with the partner hPMS2 was analysed in a yeast-2-hybrid system.
Additionally these mutations were analysed with the dominant neg-
ative mutator phenotype assay. In this system the interference of the
human MLH1 protein with the yeast mismatch repair system reveals
information about the protein function.
To determine the mutation rate, an existing yeast LYS2A14
reporter system was used. The endogenous lysine gene contains a
frameshift due to the insertion of an (A)14 run, and cells are lysine
auxotroph. Reversion to prototrophy primarily occurs through a one-
nucleotide deletion, which is caused by a defective repair system
during replication. For further characterisation of the missense
mutations, the stability of the protein was determined by Western Blot
analysis. In these studies 17 of 26 missense mutations in the hMLH1
gene were classified as pathogenic. The characterisation of 5 missense
mutations in the ATPase domain revealed only slight effects on
protein function and 4 missense mutations in the hPMS2-interaction
domain were classified as non-pathogenic.
Methods Molecular genetic studies undertaken in two large Irish
kindreds that satisfy the stringent Amsterdam criteria will be described.
Conclusion IHC as a primary molecular screening modality would
fail to identify two missense mutations. MSI analysis could poten-
tially identify a tumour that has defective DNA MMR but intact IHC
staining caused by non-truncating missense mutations. In conjunction
with a comprehensively developed family history, tumour IHC
analysis of all four MMR proteins (hMSH2, hMLH1, hMSH6 &
hPMS2) in conjunction with MSI testing is the optimum molecular
strategy to screen for Lynch Syndrome.
References
1. Lynch HT, de la Chapelle (2003) Hereditary colorectal cancer. N
Engl J Med 348:919–932.
2. Boland CR (2007) Clinical uses of microsatellite instability
testing in colorectal cancer: an ongoing challenge. J Clin Oncol
25:754–755.
Optimum clinical & molecular strategies to screen
for Lynch syndrome
Michael P. Farrell, M. J. Kennedy, D. Flannery, T. J. Boyle,
B. J. Mehigan, R. B. Stephens, S. A. White, C. B. Muldoon1,
A. J. Green, S. T. Duke2
1St. James s Hospital & Trinity College Dublin, 2National Centre for
Medical Genetics, Dublin
Background The Lynch Syndrome is a highly penetrant, autosomal
dominant, multi-system cancer disorder caused mainly by heritable
defects in the highly conserved DNA mismatch-repair (MMR) genes
hMSH2, hMLH1, hMSH6 & hPMS2 1.
Identification of a pathogenic germline mutation is extremely
important because it enables pre-symptomatic testing of family
members and structured surveillance of mutation carriers. Due to
the heterogeneity of the mutation spectrum of the MMR genes,
mutation analysis is time-consuming and expensive, therefore,
screening strategies are required to pre-select those families that
are likely to harbour a deleterious mutation. Various criteria
(Amsterdam & Bethesda) have not proved definitive for identifying
patients who may harbour a mutation. In addition to family history,
histological, immunohistochemical and molecular analysis can be
utilised to identify patients eligible for mutation analysis. Most
tumours from patients with Lynch Syndrome have a characteristic
molecular signature resulting from the involvement of defective
MMR, i.e., the presence of microsatellite instability (MSI) and/or
the absence of MMR protein expression by immunohistochemistry
(IHC) 2.
Preliminary findings from a qualitative study
of the reasons individuals decline the offer of genetic
testing for colorectal cancer
Louise A. Keogh, Belinda Mcclaren, Judith Maskiell, Clara Gaff,
John Hopper, Mark Jenkins
Key Centre for Women’s health in Society, The University
of Melbourne, Carlton, Australia
Since 1997, the Australasian Colorectal Cancer Family Registry
(ACCFR) has been studying a large number of individuals who have
Abstracts 731
123
had bowel cancer and their relatives. When a genetic mutation is
identified by the ACCFR, family members are offered the chance to
have genetic testing through a family cancer clinic to learn of their
result. We know from quantitative analysis that between 17% (before
August 2003) and 49% (after August 2003) of those offered genetic
testing decline the offer, but so far we have not known the reasons
why. We are currently following up a small group of the 120 par-
ticipants who have been offered genetic testing to explore genetic
testing decision-making in more detail. Twenty to thirty participants
will take part in a qualitative interview about genetic testing decision-
making in the area of bowel cancer. This will include 10–15 who have
declined genetic testing and 10–15 who have accepted an offer of
genetic testing. Thematic analysis will be used to determine the
reasons for accepting or declining the offer of genetic testing. Pre-
liminary findings based on interviews with decliners will be
presented. We will report why participants decline the offer of genetic
testing. A common reason given by participants for declining was the
fear of losing death and disability entitlements or becoming ineligible
for life insurance if found to carry a mutation. Individual’s perception
of their personal risk of colorectal cancer as well as their perception of
the value of knowing their mutation status consistently played a role
in decision-making.
Soft tissue sarcomas and hereditary non polyposis
colorectal cancer (HNPCC) syndrome: formulation
of an hypothesis
E. Urso1, M. Agostini1, S. Pucciarelli1, I. Mammi1, A. Viel2,
I. Maretto1, D. Nitti1
lClinica Chirurgica 2^, Dipartimento di Scienze Oncologiche
e Chirurgiche, Azienda Ospedaliera-Universita di Padova, Italy;2Oncologia Sperimentale 1^, Centro Regionale Oncologico,
IRCCS, Aviano, Italy
Background Hereditary non polyposis colorectal cancer (HNPCC)
is an inherited disorder caused by mismatch repair (MMR) gene(s)
deficiency. HNPCC predisposes to several malignances (particularly
colorectal and endometrial cancer) which not yet include soft tissue
sarcomas.
Aim To sustain the hypothesis that sarcomas could be comprised in
the spectrum of HNPCC related tumors.
Methods We report on a HNPCC patient with leiomyosarcoma of
deltoid muscle and review the literature, finding four other cases of
soft tissue tumors involving HNPCC patients.
Results Data for supporting our hypothesis are the follow: sarcomas
are rare tumours and so there is a low probability of random clustering
with colorectal and endometrial cancer; mean age at diagnosis of
sarcomas in HNPCC patients was 34 years vs. 56 years of the general
population; sporadic sarcomas are not associated to microsatellite
instability neither to loss of MMR proteins expression at immuno-
histochemistry, while all the five cases collected of HNPCC patients
with soft tissue sarcoma, presented loss of MSH2 at immunohisto-
chemistry and/or germline MSH2 mutation. Moreover, an excess of
sarcomas in MMR defective mice was also reported.
Conclusions Sarcomas could be associated to colorectal cancer and
other HNPCC related tumors as the expression of the same germline
disease. This report would encourage the scientific community to
revise the HNPCC family trees, with particular attention to soft tissue
tumors. More extensive data could confirm our hypothesis and broad
the spectrum of HNPCC related cancers.
Adenomas in Lynch syndrome: diagnostically useful?
Michael D. Walsh, Daniel D. Buchanan, Sven Arnold, Mark
Clendenning, Rhiannon Walters, John L. Hopper, Mark A. Jenkins,
Jeremy R. Jass, Joanne Young
Queensland Institute of Medical Research, Brisbane, Australia
Debate continues as to the usefulness of testing adenomas for loss of
mismatch repair (MMR) proteins to identify individuals with sus-
pected Lynch syndrome. Some studies have suggested that it is only
worthwhile testing large, proximal adenomas exhibiting severe dys-
plasia while others report disappointing ‘‘hit rates’’ even from known
mutation carriers.
We tested 77 adenomas from 49 mutation carriers arising from 38
Lynch syndrome families (14 MLH1, 20 MSH2, 3 MSH6 and 1
PMS2 mutation respectively) enrolled in the Australasian Colorectal
Cancer Family Study. The average age of diagnosis of first polyp
studied was 49.6 ± 10.6 years (30.0–79.2 years).All polyps were
tested by immunohistochemistry (IHC) for four MMR proteins
MLH1, MSH2, MSH6 and PMS2. Sixteen polyps demonstrated
normal MMR IHC (6/16 polyps came from individuals who had other
polyps which demonstrated abnormal IHC), whilst 61/77 (79.2%)
adenomas from mutation carriers showed loss of appropriate protein
expression. A subset of polyps was also assessed for evidence of
microsatellite instability (MSI) with concordance between IHC and
MSI testing in 30/32 (94%) cases.
Here was no significant difference between adenoma site with
20/23 (87%) proximal vs. 26/30 (87%) distal showing MMR defi-
ciency. Similarly, size was not a predictor of MMR deficiency with
23/31 (74%) adenomas \5 mm maximum dimension, 18/22 (82%)
5–10 mm, and 14/17 (82%) [ 10 mm MMR deficient. Of the MMR
deficient polyps, 32 showed mild dysplasia (75%), 32 showed mod-
erate dysplasia (84%) and 10 showed severe dysplasia (90%). Whilst
there was a trend to greater likelihood of loss of MMR protein with
increasing dysplasia, this was not statistically significant.
In conclusion, testing adenomas from patients from Lynch syn-
drome families is a useful alternative in cases where cancers are
unavailable since 79.2% adenomas from carriers show appropriate
loss of MMR proteins and a high level of concordant microsatellite
instability.
Patient preferences for risk management strategies
in women with Lynch syndrome
Charlotte C. Sun, Susan K. Peterson, Kristin White, Beatty Watts,
Molly Daniels, Stephanie Boyd-Rogers, Kathleen Schmeler,
Diane C. Bodurka, Karen H. Lu
University Texas M. D. Anderson Cancer Center, Houston,
Texas, USA
Background Women with Lynch syndrome must balance clinical
benefits of cancer prevention and screening strategies against poten-
tial detriments to quality of life. We assessed women’s preferences for
screening, chemoprevention, and prophylactic surgical strategies.
Methods We used the visual analog scale (VAS) and standard
gamble (SG) to assess preferences of women with Lynch syndrome.
The VAS asked women to rank strategies from 0 = worst to
100 = best. The SG asked what percentage of lifetime risk of cancer
they needed to accept each strategy. Strategies included: endometrial
screening/biopsy (GYN); (2) colonoscopy (CLSCPY); (3) combined
GYN + CLSCPY; (4) prophylactic hysterectomy/oophorectomy
(TAH/BSO); (5) prophylactic subtotal colectomy (SUBTCOL); (6)
732 Abstracts
123
combined TAH/BSO + SUBTCOL; (7) oral contraceptives (OCPs);
(8) COX-2 inhibitors (COX2); (9) combined OCPs + COX2.
Results 50 women participated (median age = 37.7 years, range
24.7–57.9). Using the VAS, the most favorable strategy was annual
combined GYN + CLSCPY (VAS = 98), followed by annual GYN,
CLSCPY, and OCPs (VAS = 90). The least preferred strategy was
combined premenopausal TAH/BSO + SUBTCOL (VAS = 28).
Postmenopausal TAH/BSO was the most favorable surgical interven-
tion (VAS = 85). If lifetime risk of cancer exceeded 40%, women
accepted postmenopausal TAH/BSO; if lifetime risk exceeded 70%,
women accepted premenopausal TAH/BSO. Women accepted
SUBTCOL if lifetime risk exceeded 75%. Compared to unaffected
women, women with a personal history of colon cancer gave less
favorable scores for COX2 (90 vs. 50, p = .001), OCP + COX2 (85 vs.
40, p = .02), CLSCPY (95 vs. 80, p = .06) but were more willing to
accept annual and biannual GYN screening at lower lifetime risk of
cancer (30% vs. 20%, p = .042; 30% vs. 10%, p = .019, respectively).
Conclusions Our data indicate that women with Lynch syndrome
had the highest preferences for combined GYN + CLSCPY screening.
Women favored all screening strategies over surgery. Reproductive
age influences preferences for TAH/BSO. Women with a personal
history of colon cancer accept GYN screening more readily than
unaffected women.
Identification of the MLH1 and MSH2 genes mutations
in Greek colorectal cancer patients
Georgia Thodi, Fostira Florentia, Sandaltzopoulos Raphael,
Yannoukakos Drakoulis
National Center of Scientific Research ‘‘Demokritos’’, Athens, Greece
Germline mutations in the mismatch repair genes, MLH1, MSH2,
MSH6 and PMS2, predispose to the Lynch syndrome. Approximately
90% of the deleterious alterations, reported in putative Lynch syn-
drome patients so far, are scattered throughout the coding regions of
the MLH1 and MSH2 genes and involve not only small aberrations
but also large genomic rearrangements. Thus, their identification
requires full sequencing of the implicated genes, as well as techniques
for the detection of gross aberrations.
Our main goal is to study the nature and frequency of the MLH1
and MSH2 pathogenic mutations encountered in Greek colorectal
cancer patients with family history indicative of Lynch syndrome in
order to facilitate the DNA-testing procedure.
We have screened twenty families selected upon modified
Amsterdam criteria or revised Bethesda guidelines for the presence of
pathogenic alterations in MLH1 and MSH2 genes using both
sequencing and multiplex ligation dependent-probe amplification.
Four point mutations, three nonsense in the exons five, seven,
thirteen of the MSH2 gene and one splice site alteration in the
intron nine of the MLH1 gene, as well as three genomic rear-
rangements, one deletion, one duplication involving exon six of the
MLH1 gene and one duplication in the exon twelve of the MSH2
gene, have been detected in eight among our fifteen Amsterdam
positive families. The deletion was characterized using long range
PCR while the duplications have not been confirmed with an
independent method, yet. The nonsense alterations located in exons
five and seven of the MSH2 gene, respectively, as well as the two
duplications have not been described elsewhere, to the best of our
knowledge.
In conclusion, the MLH1 and MSH2 mutational spectrum
observed in our patients’ cohort is quite heterogeneous while genomic
deletions/duplications seem to contribute significantly to its config-
uration.
Microsatellite instability for Lynch syndrome diagnosis
in Chile: a multicentric study
Carolina A. Rıos1, Jorge Munoz1, Leonardo Espindola2, Juan Ignacio
Vergara2, Mario Abedrapo3, Claudio Munoz3, Yamile Corredoira4,
Jaime Contreras5, Isabel Castro6, Monica Acuna1, Lucıa Cifuentes1
1Laboratorio de Epidemiologıa Genetica, Facultad de Medicina,
Universidad de Chile; 2Servicio de Cirugıa, Hospital Militar
de Santiago; 3Servicio de Cirugıa, Hospital Clınico Universidad
de Chile; 4Departamento de Anatomıa Patologica, Hospital Clınico
San Borja Arriaran, Campus Centro, Facultad de Medicina,
Uiversidad de Chile; 5Departamento de Cirugıa, Hospital Clınico San
Borja Arriaran, Campus Centro, Facultad de Medicina, Universidad
de Chile; 6Escuela de Tecnologıa Medica, Facultad de Medicina,
Universidad de Chile. Santiago, Chile
Lynch syndrome is the most common hereditary colorectal cancer
syndrome. In Chile, few studies have been developed regarding this
syndrome, although the colorectal cancer mortality rate has increased
in the last few years.
The aim of our research is to establish diagnostic criteria for
Chilean patients, considering the population characteristics, the
available data from the patients, molecular tests and the final cost-
effectiveness. To accomplish this, we have started a multicentric
study involving several hospitals from the area of Santiago, Chile.
The patients, who meet the Revised Bethesda Guidelines (RBG)
for identifying individuals at risk for HNPCC, were selected and
informed consent was obtained from all of them. Fixed and fresh
samples were collected, according to the availability, and DNA was
obtained with the proper methodology.
At the moment, 25 patients have been recruited from 4 different
hospitals. 56% of the patients meet the age criterion and only 24%
fulfil the family history criterion, probably caused by the lack of
information. The rest of the patients had synchronous or metachro-
nous colorectal cancer or tumours with MSI-high histology.
The microsatellite instability (MSI) was studied using 5 loci: Bat-
25, Bat-26, D2S123, D5S346 and D17S250. The results show 9
patients (36%) with MSI-high (with 2 or more out of 5 loci unstable).
The immunohistochemistry (IHC) for hMSH2 and hMLH1 is being
developed at the moment. All samples with MSI or altered IHC will
eventually be subjected to a BRAF V600E mutation analysis and
MSH2 and MLH1 mutation scan. (Grants: Universidad de Chile
FEBA-157, Conicyt AT-24080077).
‘‘Minor’’ mismatch repair genes involvement in genetic
predisposition to Lynch syndrome
F. Duraturo, R. Liccardo, A. Cavallo, M. De Rosa, P. Izzo
Department of Medical Biochemistry and Biotechnology,
University of Naples Federico II, Naples, Italy
Lynch syndrome is commonly associated to mutations in at least three
mismatch repair (MMR) genes: MLH1, MSH2 and MSH6. However,
mutations in these three genes do not account for all Lynch syndrome
families. Recently, it has been shown that germline mutations in
others MMR genes, PMS2 and MLH3, play a far more important role
in Lynch Syndrome than initially thought.
So far, MSH3 gene germline mutations have not been described.
In this study, a set of 63 Lynch syndrome unrelated patients, negative
for germline mutations within MSH2 and MLH1 genes, were
screened for mutations in ‘‘minor’’ MMR genes: MSH6, PMS2,
MLH3 and MSH3. Patients fulfilling the Amsterdam Criteria, show-
ing early and late onset colorectal cancer and microsatellite instable
tumors, were selected for this study. ‘‘Minor’’ MMR genes were
Abstracts 733
123
screened by denaturing high performance liquid chromatography
(DHPLC). All samples exhibiting abnormal DHPLC profiles were
analyzed by directed sequencing.
We have identified 2 frameshift mutations causing a premature
stop codon, 5 missense mutations, 7 silent mutations and 4 intronic
changes within MSH6 gene; 11 missense mutations, 4 silent muta-
tions and 7 intronic changes in PMS2 gene; 6 missense mutations, 1
frameshift mutation mutations causing a premature stop codon, 2
silent mutations and 4 intronic changes within MLH3 gene. Finally,
we have identified 4 missense mutations, 1 silent mutation and 7
intronic changes within MSH3 gene. Several patients were carriers of
at least two genomic variants of the ‘‘minor’’ genes. Thirty-four of the
65 genomic variants identified are novel, not reported in the inter-
national mutational databases.
This study gives the first evidence of MSH3 germline mutations in
Lynch Syndrome patients. Moreover, the simultaneous presence of
several mutations in different minor genes could suggest that these
genes might work together in a cooperative manner resulting in an
increased risk of cancer in Lynch Syndrome.
Prevention of endometrial tumours, a randomised
control trial of the effect of the Mirena intrauterine
system (IUS) with surveillance, versus surveillance
alone, on the development of atypical endometrial
hyperplasia (AEH) and carcinoma (EC) in women
with Lynch syndrome AGED 35-65Y
Shirley Hodgson1, Peter Sasieni2, Victoria Murday3, Yorkhill
Hospital, Glasgow; Eamonn Sheridan, Leeds General Infirmary,
Leeds
1St Georges, University of London; 2Queen Mary University
of London; 3Yorkhill Hospital, Glasgow; Eamonn Sheridan,
Leeds General Infirmary, Leeds
Introduction Women with Lynch syndrome have an increased risk
of developing endometrial cancer (up to 65% lifetime risk, 20%
before 50 years). The Mirena progestogen-releasing intra-uterine
system (IUS) greatly reduces the thickness of the endometrium.
Methods We hope to evaluate whether treatment with the Mirena for
4 years reduces the rate of detection of AEH and EC in women with
Lynch Syndrome on surveillance by annual transvaginal ultrasound
(TVS) and endometrial biopsy (EB). Secondary outcomes include
determination of the sensitivity and specificity of surveillance, the age-
related incidence of AEH and EC, the premalignant pathway to car-
cinoma and any adverse effects of surveillance and use of the IUS.
Results Unfortunately we have experienced great difficulty in
ascertaining women for the study, largely because appropriate women
within have often had hysterectomies or are eligible but unwilling to
be randomised.
Currently, 17 sites are active in the UK, with a further 7 going
through the approvals process. 17 patients have been randomised,
with a further 9 consented. Interest from non-UK sites could take the
study internationally.
Conclusion If the study demonstrates a significant reduction in the
development of endometrial neoplasia change in women with the
Mirena IUS, it will have a very substantial impact on the management
of women with Lynch Syndrome, since many such women do not
wish to have a hysterectomy, particularly during childbearing years,
but are worried that surveillance may not detect endometrial cancer
early enough for effective treatment.
The effectiveness of surveillance is still being evaluated and
women may well prefer the option of Mirena IUS if shown to be
effective. However, as the study may not be feasible, we are pro-
posing to do an observational study on all women ascertained by the
study to try and establish from this data what the diagnostic effec-
tiveness of endometrial ultrasound and biopsy is, and the effect of any
hormonal treatment.
Is uptake of genetic testing for colorectal cancer
influenced by knowledge of insurance implications?
Louise A. Keogh, David M. Studdert, Judith Maskiell, Finlay Macrae,
D. James St John, Clara Gaff, Mary Anne Young, Melissa C. Southey,
Graham G. Giles, Doreen Rosenthal, John L. Hopper, Mark A.
Jenkins
Key Centre for Women’s health in Society, The University
of Melbourne, Carlton, Australia
Objective To assess whether knowledge of insurance implications
influenced uptake of genetic testing by participants in a research study
of the causes of colorectal cancer.
Design and setting From 1999 to 2003 participants in the popula-
tion-based Victorian Colorectal Cancer Family Study were not
informed of any potential effect of genetic testing on their eligibility
for future insurance due to their participation. In 2003 the protocol
was changed when the investigators became aware that offering
participants the opportunity to learn of relevant genetic information
might affect their ability to purchase some forms of future insurance.
Since 2003 an added step was introduced to the consent procedure,
and information on the potential effect on insurance eligibility was
provided to participants.
Main outcome measure Uptake of genetic testing for mutations in
DNA mismatch repair (MMR) genes at a family cancer clinic.
Results Compared with 1999–2003, after 2003 the proportion of
participants that declined the offer of genetic testing more than
doubled from 19 to 49% (p = 0.002). This decline in uptake of
genetic testing could not be explained statistically by adjustment for
measured putative predictors.
Conclusion Knowing one has a mutation in a MMR gene is
potentially of substantial clinical importance, so the identification and
screening of people with a mutation might be a cost-effective way to
reduce the burden of colorectal cancer in the community. If indi-
viduals are choosing not to get this genetic information because of
how it will affect their eligibility for insurance, reforms to existing
insurance practices are indicated.
Utility of the immunochemical expression profile
in the identification of patients with Lynch’s syndrome
E. Alcaraz, C. Alenda, A. Paya, L. Perez, R. Jover, E. Rojas,
J. L. Soto, A. Castillejo, V. M. Barbera
Hospital General Universitario Alicante, Alicante, Spain
Background MLH1 inactivation may be observed in sporadic and
Lynch’s syndrome colorectal carcinoma (CRC). Lynch syndrome is
caused by germline mutations in the DNA mismatch repair (MMR)
genes. Sporadic CRC is caused by MLH1 promoter methylation. The
aim of this study is to evaluate the value of an immunohistochemistry
(IHC) panel in the prediction of germline MLH1 mutation in patients
with tumours that show loss of MLH1 expression.
734 Abstracts
123
Methods The study was performed in 159 CRC with loss of MLH1
IHC expression from patients of the Genetic Counselling in Cancer
Department of HGUE and from a series of non-selected CRC speci-
mens from the EPICOLON study and the HGUA. IHC analysis
for p53, p16, β-catenina, CK20, CDX-2, COX-2, CK7 and
β2-microglobulina was performed on tissue microarray (TMA).
The MLH1 methylation analysis was performed by real-time PCR
assay Methylight. V600E BRAF mutation was detected using specific
TaqMan probes by real time PCR. 48 patients were classified as
familial, 79 were classified as sporadic and 33 had not enough
information for classification.
Results There was found statistically significant association
between familiar status and IHC expression of p53 (familiar 2.6% vs.
sporadic 15.9%, p \ 0.05), p16 (familiar 100% vs. sporadic 62.5%,
p \ 0.001), CDX-2 (familiar 81.3% vs. sporadic 51.9%, p \ 0.01),
COX-2 (familiar 34.2% vs. sporadic 55.4%, p \ 0.05) and β
2-microglobulin (familiar 60% vs. sporadic 81.8, p \ 0.05). In the
regression model analysis p16 and β2-microglobulin resulted to
be independents variables that help to detect the familiar status of
CRC MLH1 negatives.
Conclusions CRC with MLH1 inactivation show a different IHC
profile according to hereditary or familiar nature. Differences founded
with the markers used in this study allow create a prediction model for
familiar or sporadic CRC which could be useful in the clinical practice.
No association between MUTYH AND MSH6 germline
mutations in 64 HNPCC patients
Verena Steinke1, Nils Rahner1, Monika Morak2, Gisela Keller3,
Hans K. Schackert4, Heike Gorgens4, Wolff Schmiegel5,
Brigitte Royer-Pokora6, Wolfgang Dietmaier7, Matthias Kloor8,
Christoph Engel9, Peter Propping1, Stefan Aretz1,
The German HNPCC Consortium
1Institute of Human Genetics, University of Bonn, Bonn, Germany;2Institute of Human Genetics, Ludwig-Maximilians-University,
Munich, Germany, and Center of Medical Genetics, Munich,
Germany; 3Department of Pathology, Munich University
of Technology, Munich, Germany; 4Department of Surgical Research,
Technische Universitat Dresden, Dresden, Germany; 5Department
of Medicine, Knappschaftskrankenhaus, Ruhr-University Bochum,
Bochum, Germany; 6Institute of Human Genetics, Heinrich Heine
University, Duesseldorf, Germany; 7Department of Pathology,
University of Regensburg, Regensburg, Germany; 8Institute
of Molecular Pathology, University of Heidelberg, Heidelberg,
Germany; 9Institute for Medical Informatics, Statistics and
Epidemiology, University of Leipzig, Leipzig, Germany
Biallelic mutations in the base excision repair (BER) gene MUTYH
are associated with multiple adenomas and an increased risk of
colorectal cancer. The heterozygote carrier frequency in the general
population is approximately 1–2%. Gu et al. (2002) reported on an
interaction of MUTYH and mismatch repair (MMR) proteins. They
showed that the MSH2/MSH6 complex enhances the binding affinity
to the mismatched DNA substrate and the glycosylase activities of
MUTYH.
Mutations in MMR genes are known to cause hereditary non-
polyposis colorectal cancer (HNPCC/Lynch syndrome), an autosomal
dominant tumor predisposition syndrome. Based on the reported
interaction of the MMR complex and the base excision repair protein
MUTYH, it was hypothesised that MUTYH mutations serve as phe-
notypical modifiers in HNPCC families. Recently, a significantly
higher frequency of heterozygosity for MUTYH mutations among
MSH6 mutation carriers was reported.
We examined 64 MSH6 mutation carriers (42 truncating mutations,
19 missense mutations and 3 silent mutations) of the German HNPCC
Consortium for MUTYH mutations by sequencing the complete
coding region of the MUTYH gene. We identified monoallelic MU-
TYH mutations in two of the 64 patients (3.1%): the truncating
MUTYH mutation c.247C [ T;p.Arg83X was found in a patient
harbouring a pathogenic MSH6 mutation and the missense mutation
c.502C [ T;p.Arg168Cys was identified in a carrier of a MSH6 silent
mutation of unknown functional relevance. No biallelic MUTYH
mutation carrier was found. The frequency of MUTYH mutations was
not significantly higher than in healthy controls, neither in the whole
patient group (p = 0.30) nor in different subgroups regarding muta-
tion type. Our results do not support an association between MSH6 and
heterozygosity for MUTYH mutations.
The study was supported by the Deutsche Krebshilfe.
Lynch syndrome in Rosario, Argentina: 3 years
experience
Enrique Spirandelli1, Sergio Chialina1, Florencia Spirandelli2,
Fernando Serra2, Ariel Naves3, Claudio Settecase2, Juan Carlos
Moreno2, Gustavo Castellani2, Gustavo Botti2, Ana Maria Moriena2,
Luciana Spirandelli2
2Integrantes del Servicio de Coloproctologia del Hospital Espanol-
.1STEM SRL; 3Instituto de Histopatologia jefe del servicio de
Coloproctologia Hospital Espanol de Rosario
The Lynch Syndrome is characterized by the development of colo-
rectal cancer, endometrial cancer and other associated cancers. It is
caused by a mutation in one of the mismatch repair genes: mainly in
MLH1 and MSH2 but also in other genes, such as MSH6 or PMS2. In
Rosario city, Argentina, which has a population of 1.200.000 inhab-
itants we have established since 2005 a multidisciplinary group
named ACETIEHR for the diagnosis and management of patients
with hereditary cancer such as the Lynch Syndrome.
Materials and methods Between September 2005 and December
2008 we have already identified 71 probands (male: 54%, female:
46%) fulfilling the revised Bethesda guidelines. Mean age at diag-
nosis 52.9 (range 17–81 years). We found extracolonic tumors in 57
probands and 86 family members (most frequent 31 in females and 55
in males). All the probands and their relatives received genetic
counseling.
We have already performed 29 microsatellite instability (MSI)t-
ests using 5 markers (BAT-25, BAT-26, DS17250, D5S346,
D2S123). We are also setting up immunohistochemistry which
combined with MSI profiling is the current method of identifying the
gene to be sequenced.
Results We found 9 tumors with positive MSI. In 3 probands
MSI(+)who underwent molecular sequencing, a pathologic mutation
was found: 2 in MSH2 gene (c1864 C-[T exon 12 and c1224 T-[A
exon 7) and 1 in MLH1 (c1852_1854 del AAG exon 16).Ten persons
at risk were tested, and of these, 7 had the mutation and 3 did not.
Discussion We estimate that in Rosario city there are 500–2000
mutation carriers in risk of developing colorectal cancer and other
cancers associated with the Lynch Syndrome that could be early
detected in order to adopt medical prevention measures. Because of
the accumulating evidence that MSI is a predictive factor for response
to 5FU-based chemotherapy, our group considers that MSI status will
be a relevant prognostic factor for all colorectal cancers.
Abstracts 735
123
A prospective single blinded randomized trial
of polyethylene glycol-electrolyte solution vs. sodium
phosphate as a bowel preparation for colonoscopy
in Lynch syndrome gene carriers
M. W. J. van Vugt-van Pinxteren, M. H. van Kouwen,
M. G. H. van Oijen, F. M. Nagengast
Department of Gastroenterology, UMC St. Radboud Nijmegen,
The Netherlands
Background Lynch syndrome gene carriers undergo regular sur-
veillance colonoscopies. Polyethylene glycol-electrolyte solution
(PEG) is routinely prescribed for bowel cleansing but often poorly
tolerated by these patients. Sodium phosphate (NaP) may be an
effective alternative.
Aim To randomly compare the effects of bowel preparation on
colonic cleansing and patients’ acceptance.
Methods During a 1 year inclusion period Lynch syndrome patients,
who previously underwent a colonoscopy were invited to participate.
Patients were randomly assigned to either PEG or NaP and asked to
fill in a questionnaire about preparation tolerability and future pref-
erences prior to and 1 week after the preparation for colonoscopy.
The endoscopist (blinded for the preparation) filled out a report about
the colon cleansing.
Results A total of 125 carriers were included in the study. Nine
(7%) were excluded because of missing data. The remaining 116
patients (M/F 58/58, mean age 50 ± 30 years) were included in the
statistical analysis. Of those, 53 used PEG and 63 NaP. Baseline
characteristics did not differ between groups. Of the patients using
PEG, 13% found the preparation almost intolerable, in contrast to 8%
of those using NaP (p = 0.005). Future preparation preferences were
18% for PEG, 51% for NaP, 27% did not have a preference (of 4% no
data). The colonoscopy was poorly tolerated in 28% of individuals
using PEG and in 25% of the NaP participants (p = 0.963). The
endoscopist reported a good to excellent clean colon in 83% of the
patients on PEG and in 71% of those on NaP (p = 0.096). The pro-
portion of colonoscopies with introduction into the cecum within
25 min did not differ between groups: 68% PEG and 72% NaP
(p = 0.645).
Conclusion Lynch syndrome carriers tolerated NaP better and pre-
ferred this formula for bowel preparation. Colon cleansing was
suboptimal by both treatments with a tendency towards a cleaner
colon with PEG.
Capsule endoscopy for the small bowel in juvenile
polyposis syndrome
O. Will, A. J. Postgate, C. H. Frase, A. Fitzpatrick, R. K. S. Phillips,
S. K. Clark
St. Mark’s Hospital, Middlesex, UK
Introduction Juvenile polyposis syndrome (JPS) is one of the hamar-
tomatous polyposis syndromes and demonstrates phenotypic hetero-
geneity. All patients with JPS develop colorectal polyps and are at risk
of colorectal cancer. Small bowel involvement in JPS is variably
described. Small intestinal cancer is reported but is rare and currently
there is no evidence-based protocol for small intestinal surveillance.
Aim This study investigates the utility of capsule endoscopy (CE) in
patients with JPS.
Methods Ten patients with JPS underwent CE. Medical records
were reviewed to characterise the genotype. Conventional upper and
lower gastrointestinal endoscopy were used to characterise the phe-
notype of the rest of the bowel.
Results Two patients had small bowel polyps beyond the range of
oesophagogastroduodenoscopy (OGD) identified at CE: a 6 mm ileal
polyp in one and 10 and 6 mm ileal polyps in the second. In addition
duodenal polyps were detected in a third patient at CE. Three further
patients had previously documented duodenal polyps at surveillance
OGD. A SMAD4 mutation was identified in 7 patients and there was
no obvious association with gastric/small bowel polyposis. Patients
reported CE as comfortable and convenient.
Conclusions CE provided information additional to conventional
endoscopy and was well-tolerated. However no lesions requiring
clinical intervention were identified and polyp numbers were small.
CE may be used as a non-invasive method of investigating the small
bowel JPS, but there is no evidence to support routine surveillance in
colonic-confined JPS.
A novel mutation in MUTYH: associated polyposis
(MAP) syndrome
Florentia Fostira1, Christos Panopoulos2, Anna Efraimidis2,
Drakoulis Yannoukakos1
1N.C.S.R ‘‘Demokritos’’, Molecular Diagnostics Laboratory,
R-RP Institute, Athens, Greece; 2Oncology Anticancer Hospital
‘Agios Sabbas’, 2nd Department of Medical Oncology, Athens,
Greece
MUTYH-Associated Polyposis (MAP) syndrome is inherited as a
recessive trait and is characterized by the intrinsic phenotypic fea-
ture of adenomas present at the individual’s colorectum. In the case
of non-surgical removal of these polyps, adenoma will transform to
carcinoma. Colorectal tumours from mutation carriers display an
excess of somatic mutations, specifically G:C to T:A transversions.
The risk of colorectal cancer among biallelic mutations carriers
seems to be close to 100% by the age of 60. Four families meeting
the MAP phenotypic criteria were studied. Genomic DNA was
purified from peripheral blood leukocytes following standard chlo-
roform extraction. The complete coding sequences of the MUTYH-
gene, including splice junctions, were amplified by Polymerase
Chain Reaction (PCR) and electrophorized in an ABI Prism� 310
Genetic Analyzer. Analysis of the MUTYH gene revealed five
different germline mutations, one of which novel and never char-
acterized in respects of its pathogenicity. A mutation has been
identified on exon 7 of the MUTYH gene at a homozygous state, in
an individual with less than one hundred colorectal polyps at the age
of forty. Another patient diagnosed with colorectal cancer at the age
of 34 carried two maternally and one paternally inherited mutation,
located on exon 14. A milder MAP phenotype characterized an
individual, who was diagnosed with less than one hundred polyps at
the age of 69. Genetic testing revealed that the patient was com-
pound heterozygote for two missense variants located on exons 8
and 12 of the MUTYH gene. A novel mutation located on intron 6
of the MUTYH gene has been identified in two families. Our
findings confirm previous observations highlighting the necessity for
genetic testing of MUTYH in patients with MAP phenotypic fea-
tures, as well as his/her extended family, in order to determine the
phenotyping expression of each mutation and propose the appro-
priate clinical surveillance, which includes biennial colonoscopy
after the age of 30. In specialized cases surgical intervention might
be essential.
736 Abstracts
123
APC or MUTYH mutations account for the majority
of clinically well characterized families with familial
adenomatous polyposis and attenuated familial
adenomatous polyposis phenotype and patients
with more than 30 adenomas
Bruno Filipe1, Celia Baltazar1, Cristina Albuquerque1,
Sofia Fragoso1, Pedro Lage2, Ines Vitoriano1, Susana Mao de Ferro2,
Isabel Claro2, Paula Rodrigues2, Paulo Fidalgo2, Marılia Cravo2,
Carlos Nobre Leitao2
1Centro de Investigacao de Patobiologia Molecular (CIPM), Instituto
Portugues de Oncologia de Lisboa de Francisco Gentil, EPE,
Lisbon, Portugal; 2Department of Gastroenterology, Instituto
Portugues de Oncologia de Lisboa de Francisco Gentil, EPE, Lisbon,
Portugal
Patients presenting familial adenomatous polyposis (FAP), attenu-
ated FAP (AFAP) or multiple colorectal adenomas (MCRA)
phenotype are clinically difficult to distinguish. We aimed to eval-
uate the contribution of APC and MUTYH germline mutations for
107 clinically well characterized patients with FAP related pheno-
type, classified according with the recent guidelines for the clinical
management of FAP. Patients were stratified into 5 groups: FAP,
AFAP, MCRA (10–99 colorectal adenomas) without familial history
of colorectal cancer or few adenomas (FH), MCRA (10–99) with
FH, MCRA (3–9) with FH. Most FAP patients presented APC
mutations (38/43; 81%) and few showed MUTYH mutations (2/46;
4%), whereas AFAP patients presented a lower APC but a higher
MUTYH mutation frequency (5/16; 31% vs. 8/20; 40%). MCRA
patients did not present APC mutations but showed distinct
MUTYH mutation frequencies: 0, 2/14 (14%) and 7/13 (54%),
respectively for 3–9, 10–29 and 30–99 adenomas (P = 0.033). APC
large deletions and expression deficiency in mutation negative
patients suggest that both defects may account for a fraction of these
cases. The remaining cases may represent a new clinical entity. We
validate the recently proposed guidelines in our patient’s cohort and
show that APC or MUTYH germline defects are responsible for the
majority of clinically well characterized families with FAP, AFAP
phenotype and patients with more than 30 colorectal adenomas. The
different mutation frequencies among the five groups, specially
according with the number of adenomas, points out for the impor-
tance of excellent clinical characterization and colonoscopy
techniques for the management of FAP related phenotypes.
Adrenal incidentalomas in FAP patients
A. Hansmann, O. C. C. Will, R. K. S. Phillips, F. F. Palazzo,
K. Meeran, M. Marshall, S. K. Clark
The Polyposis Registry, St. Mark’s Hospital, Harrow, United
Kingdom Department of Radiology, St. Mark’s Hospital, Harrow,
United Kingdom; Department of Endocrine Surgery, Hammersmith
Hospital, London, United Kingdom; Department of Endocrinology,
Hammersmith Hospital, London, United Kingdom
Aim Adrenal incidentaloma (AI) is often diagnosed in patients with
Familial Adenomatous Polyposis (FAP), since they frequently undergo
cross-sectional imaging of the abdomen, and also have a higher inci-
dence of AI than the general population. This study investigates the
natural history of AI in FAP, and suggests a management protocol.
Patients and methods An original cohort of 14 FAP patients with
AI, identified 12 years ago in a prospective study, was followed up
clinically and radiologically. A further group of 16 FAP patients with
AI was also identified. All had AI [ 1 cm in size. For both cohorts,
characteristics of patients (genotype, age at diagnosis, concomitant
diagnoses) and incidentaloma (size, laterality, rate of growth, out-
come) are described.
Results Overall, three of 30 patients underwent adrenalectomy; one
had phaeochromocytoma and another an adenoma of borderline
malignancy. A further three lesions were radiologically suspicious for
malignancy at time of diagnosis, one in a patient who was unfit for
surgery but died of non-adrenal causes after more than 9 years. None
of the lesions radiologically benign at diagnosis showed an aggressive
course, but one patient required referral for surgery after 12 years of
follow-up due to slow size increase. There were no associations with
genotype.
Conclusions FAP-associated AI warrants long-term follow-up.
While the natural history is similar to that of sporadically-occurring
lesions, these patients have concomitant FAP-associated manifesta-
tions under regular radiological surveillance, and we suggest a
management protocol tailored accordingly.
Audiology in familial adenomatous polyposis:
do you hear what I hear?
Margaret O’Malley, James Church, Lisa LaGuardia, Richard Naugle,
Cynthia Gensur, Jeff Hammel, Carol A. Burke
Digestive Disease Institute, Sanford R. Weiss, MD Center
for Hereditary Colorectal Neoplasia, Cleveland Clinic, Department
of Colorectal Surgery, Department of Psychology, Department
of Audiology, Department of Gastroenterology, Cleveland,
Ohio, USA
Introduction The APC protein has an important role in maintaining
function of microtubules in the ear that play a significant part in the
mechanism of hearing. Preliminary data suggests that the APC protein
is associated with an increased incidence of abnormal hearing which
may affect intellectual function. We sought to assess the hearing
among patients with FAP (Familial Adenomatous Polyposis).
Methods Patients with FAP were recruited for an IRB-approved
study assessing hearing and intelligence. Hearing was tested by pure
tone air conduction audiometry, less than or equal to 30 db at 3/4 of the
following frequencies (500, 1000, 2000 and 4000 Hz). Subjects were
then administered the Kaufman Brief Intelligence Test (KBIT-2). We
then analyzed the proportion of individuals with an abnormal audi-
ometry as compared to age and gender adjusted normalized hearing
standards.
Results 44 patients were recruited from 42 families. Subjects
included 22 men with a mean age of 42 years. When compared to
normalized hearing standards, 19 (43.2%) of the 44 patients failed to
meet the standard normal range. Audiologic abnormalities showed
unilateral hearing impairment was documented in 6 patients, bilateral
impairment in 13. Of these patients 63% were impaired at a single
frequency; the other 37% were at multiple frequencies. 59% of
patients showed right hearing impairment with the highest deficit
(35%) at 2000 Hz. 71% of patients had left sided impairment with the
greatest number (32%) at 4000 Hz.
Conclusion A large subset of our sample of FAP patients (43.2%)
had abnormal audiologic results when compared to the normalized
standard. Differences in IQ scores for patients with and without
audiologic abnormalities are not statistically significant, suggesting
that these results do not reflect an association between hearing and
intellectual functioning in our sample.
Abstracts 737
123
Cognitive function in FAP: anyone out there listening?
Margaret O’Malley, James Church, Lisa LaGuardia, Richard Naugle,
Cynthia Gensur, Jeff Hammel, Carol A. Burke
Digestive Disease Institute, Sanford R. Weiss, MD Center
for Hereditary Colorectal Neoplasia, Cleveland Clinic, Department
of Colorectal Surgery, Department of Psychology, Department
of Audiology, Department of Gastroenterology, Cleveland,
Ohio, USA
Introduction Preliminary data suggests the APC protein is critical for
microtubule dependent pathways in the cochlea and may be important
in cognition. Abnormal audiometrics have been documented in FAP.
We studied cognitive function among patients with FAP.
Methods FAP patients were recruited fora study assessing intelli-
gence using the Kaufman Brief Intelligence Test (KBIT-2), which
provides Verbal, Nonverbal and Composite IQs. The KBIT-2 was
administered and scored by individuals experienced in administration
of psychometric measures. Mean scores were analyzed and compared
to standard normal ranges.
Results 44 Subjects from 42 families (22 men), mean age of 42 years
were included. KBIT-2 Composite IQ score was 98.4 ± 12.4 (95% CI
94.5–102.3) which is within the average range of 90–109. 27% of patients
scored below average (less than 90) and 15% scored above average
(greater than 109), not a significant imbalance (sign test p = 0.33).
Nonverbal IQ scores show no difference from average, mean = 100.5;
24% scored below and 27% scored above average. Verbal mean score
was 95.5 ± 12.0 (95% CI 91.7–99.2) significantly lower than average
(one-sample T-test P = 0.020). There is an imbalance among patients
with 27% below and 7% above the average range and a tendency toward
lower than average scores (sign test P = 0.06). The mean number of
points by which Nonverbal IQ exceeded Verbal IQ was 5.0 ± 12.5 (95%
CI 1.1–9.0) (one-sample T-test P = 0.013). The non-verbal score
exceeded the verbal score for 27 patients (65.9%), while the verbal score
was larger for only 10 patients (24.4%) (Sign test P = 0.008).
Conclusion Composite IQ scores suggest that FAP patients do not
have lower IQ than the general population. However the verbal scores
of FAP patients which are dependent on hearing are significantly
lower than average and may reflect abnormal audiometrics or other
effects of the APC mutation on cognitive function.
Your patient information website: how good is it?
Ramawad Soobrah, Anand Patel, Sue Clark
St Mark’s Hospital, Harrow, Middlesex, UK
Aim Although the Internet has greatly improved access to health
information for patients, experts have raised concerns about the overall
quality of those websites. This study was designed to evaluate the
accessibility, quality and reliability of information about familial ade-
nomatous polyposis (FAP) on the Web. The secondary aim was to
evaluate the websites of InSiGHT member institutions (IMI) separately.
Method We searched for the keywords ‘‘familial adenomatous
polyposis’’ using the three most popular search engines (Google,
Yahoo and MSN). The search was restricted using the ‘‘English
language’’ and ‘‘exact phrase’’ settings and we looked at the first 50
websites only. The LIDA tool (an online validated instrument for
health care websites) was used to assess their accessibility, usability
and reliability. LIDA scores can be classified as being high [90%),
medium (\90%, [50%) or low (\50%). The readability of each
document was assessed using the Flesch reading ease score (FRES).
A FRES score of 60–69 represents a standard readability level.
Result Of the 150 possible sites, only 55 were analysed because of
repetitions (52), irrelevant content (21) or inaccessible links (22). The
mean LIDA score was 61.9% (SD = 10) and mean FRES score was
35.2 (SD = 16). Only seven IMI websites were identified in the list
and their mean LIDA and FRES scores were 67.9% (SD = 8.8) and
47.3 (SD = 7.8) respectively. The mean reliability scores of all
websites compared to IMI websites were 37.4% (SD = 16.7) and
56.1% (SD = 9.2) respectively.
Conclusion Overall, information available for patients about FAP
on the internet is difficult to access, and of poor quality. Although the
mean LIDA and FRES scores of IMI websites tend to be higher, the
readability of their contents remain poor and they do not necessarily
appear among the top search results. Hence the need to develop clear,
easily accessible and authoritative resource for FAP patients and their
relatives.
The role of common MUTYH gene mutations
in breast cancer appears insignificant
Astrid A. Out1, Ivonne J. H. M. van Minderhout1, Carli M. Tops1,
Maartje Nielsen1, Marjan M. Weijs1, Martijn H. Breuning1,
Hans F. A. Vasen2,4, Peter Devilee1,3, Frederik J. Hes1
1Center for Human and Clinical Genetics, 2Department
of Gastroenterology, 3Department of Pathology, Leiden University
Medical Center, 4The Netherlands Foundation for the Detection
of Hereditary Tumours, Leiden, The Netherlands
Previously, we described a significantly higher frequency of breast
cancer in female bi-allelic MUTYH mutation carriers as compared to
the Dutch population (4 out of 22, Standardized Morbidity
Ratio = 3.75). Accordingly, we further investigated the possible role of
constitutional MUTYH mutations in the development of breast cancer.
A Dutch population based cohort of 1219 breast cancer patients, 476
familial breast cancer patients and 1148 controls were genotyped by
Taqman� assays for the 3 most common Dutch mutations, Y179C,
G396D and P405L, the unclassified variant R309C and reported benign
variant S515F (previously annotated as Y165C, G382D, P391L, R295C
and S501F). Additionally, direct sequencing of the whole coding part of
the MUTYH gene was performed in 306 patients. Genotyping showed
2.2, 2.5 and 1.7% heterozygote carriers for one of the three mutations
among patients, familial patients and controls respectively, giving no
significant difference. Variation in the frequencies of R309C and S515F
was also not-significant between groups. Remarkable was a doubled
frequency of G396D among familial patients as compared to controls.
Sequencing revealed 3 coding and 6 intronic rare unclassified variants,
without yet clues for pathogenicity. Overall, no bi-allelic mutation
carriers were found. In conclusion, these findings could not confirm an
association of MUTYH variants with breast cancer, although G396D
may be very weakly associated.
Detecting somatic mosaicism of the APC gene
with high resolution melting curve analysis
A. A. Out1, I. J. H. M. van Minderhout1, A. C. Schaap1,
R. Louiszoon1, E. C. Bik1, C. M. Tops1, R. H. A. M. Vossen1,
N. van der Stoep1, E. Bakker1, H. F. A. Vasen2,4, H. Morreau3,
P. Devilee1,3, F. J. Hes1
1Center for Human and Clinical Genetics, 2Department
of Gastroenterology, 3Department of Pathology, Leiden University
Medical Center, 4The Netherlands Foundation for the Detection
of Hereditary Tumours, Leiden, The Netherlands
738 Abstracts
123
Given the relatively high frequency of de novo APC mutations in
polyposis patients, a substantial proportion of mosaic APC mutations
can be expected. In a previous study 10 mosaic cases were identified
among 242 APC mutation carriers (4%) using direct sequencing, PTT
and DGGE. Scanning APC and MUTYH negative polyposis patients
with a uniform sensitive method may reveal more mosaic cases. From
208 patients, leukocyte derived DNA was scanned with High Reso-
lution Melting curve analysis (HR-MCA) for mutations in APC exons
4–6, 8–12, 14 and part of exon 5. To test the sensitivity for detection of
mosaic mutations, DNA samples with different heterozygous muta-
tions were diluted with wild-type DNA to create a range of allelic
percentages. Samples with aberrant melting curves were analyzed by
direct sequencing analysis and variants were additionally quantified by
pyrosequencing. The median detectable level of mosaicism in diluted
samples was 6%, which varied between different amplicons and
mutations (2–25%). So far, one mosaic APC mutation carrier was
detected. A C.1958+1G[T change in exon 14 was observed, showing a
minimal elevated T-peak on direct sequencing, whereas HR-MCA and
pyrosequencing analyses showed a 15–20% allele frequency. Fur-
thermore, 6 previously undetected heterozygous pathogenic mutations
were found. These results prove that HR-MCA is a promising and
sensitive method for screening and quantification of mosaic mutations.
However, the minimal detectable frequency is variable. Our aim is to
further optimize and complete the analysis for the whole APC gene, to
screen further polyposis patients and to set up mutation analysis in
formalin fixed paraffin embedded (FFPE) polyp tissue.
The MUTYH gene variant database
Astrid A. Out1, Carli M. J. Tops1, Maartje Nielsen1, Marjan M.
Weiss1, Hans F. A. Vasen2,3, Johan T. den Dunnen1, Stefan Aretz4,
Julian R. Sampson5, Peter Devilee1, Frederik J. Hes1.
1Center for Human and Clinical Genetics, 2Department
of Gastroenterology, Leiden University Medical Center,3The Netherlands Foundation for the Detection of Hereditary
Tumours, Leiden, The Netherlands, 4Institute of Human Genetics,
University of Bonn, Bonn, Germany; 5Institute of Medical Genetics,
School of Medicine, Cardiff University, Cardiff, Wales, United
Kingdom
The MUTYH gene encodes a DNA glycosylase involved in base
excision repair. A bi-allelic defect in this gene, leading to accumu-
lating somatic G to T transversions in genes like TP53, APC and
KRAS, is associated with colorectal polyposis and cancer. Whereas
much is known about the two most frequent mutations Y179C and
G396D (previously annotated as Y165C and G382D), the effect of
uncommon variants is less well known. Especially, the very rare and
unreported unclassified variants remain a holdup in molecular diag-
nostics. The open access MUTYH gene variant database (www.
lovd.nl/MUTYH) aims to supply a role in internationally collecting
and sharing of useful variant data combined with available phenotypic
information. The current content consists of data extracted from
published literature and an increasing number of unpublished variant
records submitted by different international institutes. Submitted
variants will become public after curation and annotation conform to
current mutation nomenclature (reported nomenclature also listed).
International submission could be especially helpful to catalog and
exchange information on rare variants, which would be impossible
through published literature. This locus-specific database (LSDB), in
the Leiden Open (source) Variation Database (LOVD) format, is easy
to consult and to submit information to. Furthermore, it is maintained
following the recommendations of the Human Variome Project
(HVP). With our own efforts, available and future collaborations, we
aim to maintain an up-to-date online resource of MUTYH data,
valuable for researchers and clinicians in the field.
Laparoscopic surgery in patients with colonic
polyposis syndromes
Francisco Lopez-Kostner, C. Heine, P. Medina, U. Kronberg,
C. Wainstein
Colorectal Surgery Unit, Clınica las Condes. Santiago, Chile
Background Laparoscopic colorectal surgery has been established
as a safe procedure for patients with cancer and diverticular disease of
the colon. Patients with colonics polyposis syndromes have a special
condition determined by the magnitude of the procedure (total
colectomy or proctocolectomy) and because most of the time this is a
profilactic surgery (preventing the development of cancer).
Aim To analyze results of laparoscopic surgery in patients with
colonic polyposis syndromes.
Materials and methods Data where obtained from the prospective
database of laparoscopic colorectal surgery. We selected all patients
operated on by colon polyposic syndromes (Familial adenomatous
polyposis, Familial attenuated adenomatous polyposis and Mixed
polyposis) between years 1998 and 2008.
Results In these period where operated on 25 patients with colonic
polyposis syndromes. Sixteen patients had classical familial ade-
nomatous polyposis (FAP), 9 had attenuated familial adenomatous
polyposis (AFAP) and one had mixed polyposis. The mean age was
41 years presenting a bimodal distribution according the type of
disease (average 33 years in PAF and 55 years in APAF). Fifty-six
percent were woman. Total colectomy with ileorectal anastomosis
was performed in 18 patients (72%), and restorative proctocolectomy
(ileal pelvic reservoir) in seven (28%). The mean operating time was
270 min (r: 180–400). The conversion rate was 5% (one patient) due
to anatomical difficulties. Liquid diet was started at 36 h and the
average hospital stay was 5.7 days. Complication rate was 10% (two
patients) and there was no operative mortality.
Conclusions The laparoscopic access is a feasible and a safe alter-
native for patients with colonic polyposis syndromes.
Computed tomographic colonography to assess
perioperative colorectal polyp burden in familial
adenomatous polyposis
Margaret O’Malley, James Church, Lisa LaGuardia, Jon D. Vogel,
Grace Cheah, Mark Baker, Carol A. Burke
Digestive Disease Institute, Sanford R. Weiss, MD Center
for Hereditary Colorectal Neoplasia, Cleveland Clinic, Department
of Colorectal Surgery, Department of Radiology, Department
of Gastroenterology, Cleveland, Ohio, USA
Background CT Colonography (CTC) efficacy to assess timing for
colectomy in FAP is unknown.
Methods FAP patients scheduled for primary colon surgery age
[13 years were eligible. CTC and colonoscopy were performed day
of surgery by operators blinded to results of the other study. Size and
number of colorectal polyps on CTC and colonoscopy were com-
pared. Pathologic review of the surgical specimen was performed.
Findings documented include ranges of colon and rectal polyp
number, size of largest polyp, and potential mass lesions.
Abstracts 739
123
Results Ten patients, 7 male, were enrolled (mean age 32). All
subjects had 100–1000 polyps with largest mean polyp size of 22 mm
on pathology (17.5 mm on CTC and 18.7 mm on colonoscopy). One
patient had a rectal and colon cancer detected on all 3 modalities.
CTC and colonoscopy agreed on colon polyp number in 70% but in
20% CTC count was lower than colonoscopy. The respective agree-
ment on rectal polyp number was 60%, while CTC underestimated
rectal polyp count in 40%. The number of colon polyps on CTC
agreed with pathology in 50%, but was lower in 50%. Colonoscopy
agreed with pathology on 60% but was lower in 40%. 3 patients had
proctocolectomy: colonoscopy agreed with pathology on rectal polyp
burden in 75%, CTC suggested a lower burden in 75%.
Extracolonic findings reported on CTC in 40%; one possible
pancreatic tumor, 2 patients with pulmonary nodules, and one adrenal
mass removed during surgery and confirmed adenoma. Follow up CT
ruled out a pancreatic abnormality, and pulmonary nodules were
found to be calcified granulomas.
Conclusions CTC underestimates colon and rectal polyp burden up
to 40% in FAP patients compared to colonoscopy. Both modalities
underestimate colorectal polyp burden compared to pathology but
don’t miss significant lesions. Extracolonic findings are common,
none impacted FAP surgery.
APC alternative splicing as responsible for phenotypic
variability in familial adenomatous polyposis
S. Gonzalez1, M. Menendez1, I. Blanco2, A. Obrador-Hevia3,
C. Lazaro1, G. Capella1
1Translational Research Laboratory, Catalan Institute of Oncology,2Genetic Counselling Unit, Catalan Institute of Oncology,3Biology of Cancer Group, University of Balearic Island
Familial Adenomatous polyposis (FAP) is a dominantly inherited
colorectal tumour predisposition syndrome that results from germ-line
mutations in the Adenomatous Polyposis Coli (APC) gene. FAP
shows substantial phenotypic variability: classical polyposis patients
develop more than 100 colorectal adenomas, whereas those with
attenuated polyposis (AFAP) have fewer than 100 adenomas usually
associated with late onset. Abnormalities of pre-mRNA splicing are
increasingly recognized as an important mechanism through which
gene mutations cause disease and may affect disease expression,
leading to phenotypic variability.
The aim of the study was to elucidate the molecular basis of the
phenotypic variability of one FAP and one AFAP families harbouring
the same c.834+1G[A APC mutation.
RNA-based studies were performed in six members of family A
characterized by the presence of thyroid cancer in most cases, desmoids
tumours and a classical FAP phenotype and in eight members of family
B, characterized by three generations of affected members displaying
AFAP phenotype. Eleven healthy controls were also included. Lym-
phocytes of two carriers per family were cultured and treated with
puromycin, a translational inhibitor, before RNA extraction.
Family A: In affected members RNA analyses of exons 5–9 showed
the appearance of two APC isoforms (WT, and a 11 bp frameshift
deletion that will generate a truncated protein (p.Asn276PhefsX8).
Family B: Affected relatives harbouran additional intronic change
c.730-29A[T in APC gene, that does not predict any splicing aberra-
tion. These relatives displayed a third transcript (321 bp) with an
in-frame exon 7 skipping (p.Arg244_Gln278del). Surprisingly, this
transcript also appears in family A after puromycin-treatment.
The c.834+1G[A APC mutation generates a complex pattern of
mRNA splicing. Our findings suggest that the presence of c.730-
29A[T in family B is associated with increased levels of transcripts
lacking exon 7. Differences in mRNA processing may modify phe-
notypes associates with APC gene mutations.
Haplotype analysis and age estimation of the Y165C
and G382D mutations in German MAP patients
Stefan Aretz, Stefanie Vogt, Daniela Tavian, Astrid Kaufmann,
Dietlinde Stienen, Markus M. Noethen, Sven Cichon, Waltraut Friedl,
and Roberto Colombo
Institute of Human Genetics, Bonn, Germany
Background In Caucasian MAP patients, the combined allele fre-
quency of the mutations Y165C and G382D ranges between 50 and
82%, while these mutations have not been identified in Asian popu-
lations, supporting the hypothesis of a founder effect that occurred in
the history of Caucasians.
Methods We genotyped 5 intragenic SNPs and 10 gene-flanking
STRPs in 32 unrelated German MAP patients who were homozygous
or compound-heterozygous for the Y165C and G382D mutations. To
investigate the natural history of the two common MUTYH alleles,
we calculated their apparent age in generations (g) by a set of pop-
ulation-genetic algorithms and under different assumptions on the
spread and molecular evolution of the founder haplotypes. Results. A
fully conserved intragenic SNP haplotype and two highly conserved
core STRP haplotypes were identified in 63% of mutation-bearing
chromosomes. An ancestral haplotype could be identified which
occurs at significantly increased frequency in the Y165C and G382D
chromosomes. According to parametric and Bayesian analysis of
linkage disequilibrium’s decay over time, the age of the most com-
mon recent ancestors of the two MUTYH mutations was estimated as
62–87 g (overall 95% CI 49–120 g) and 136–184 g (overall 95% CI
105–206 g) for Y165C and G382D, respectively.
Conclusions Our results suggest that the Y165C and G382D chro-
mosomes sampled in German MAP patients derive from two
ancestors that lived between the sixth century B.C. and the third
century A.D., and between the middle of the third millennium B.C.
and sixteenth century B.C., respectively. Comparative haplotype
studies on chromosomes bearing the same MUTYH mutations in
different populations will refine our knowledge of their origin and
spread into the European continent. The demonstration of founder
effects has implications for MUTYH mutation analysis in polyposis
patients of different ethnic origin.
The study was supported by the German Cancer Aid (Deutsche
Krebshilfe) and the CARIPLO Foundation.
‘‘High Risk’’ clinic for hereditary colorectal neoplasia:
a focus for patient care and an opportunity for clinical
research
Lisa LaGuardia, Margaret O’Malley, Jon D. Vogel, Brandie Leach,
Carol Burke, Matthew Kalady, James Church
Patients with Hereditary Colorectal Neoplasia and their families need
specialized care to plan for appropriate surveillance and to ensure that
they receive the most favorable treatment. This involves coordinating
multidisciplinary appointments on the same day to minimize incon-
venience to patients. We have established a special ‘‘High Risk’’ clinic
for these patients and their families. In this study we are reporting our
activity for the last 5 years.
740 Abstracts
123
Methods Initially the clinic ran one morning a month but has grown
in the last 2 years adding another half day session. Requests for
appointments were triaged by Registry Coordinators. Patients with
syndromes of Hereditary Colorectal Neoplasia were eligible for this
clinic if the necessary appointments included multiple physicians. The
Clinic is staffed by one of three colorectal surgeons, one gastroen-
terologist, one genetic counselor, one hepatobiliary/upper GI surgeon
and often by a clinical geneticist.
Results From January 2004 to November 2008 there have been 440
patient visits, 68 colonoscopies, 180 flexible sigmoidoscopies, and
226 EGD’s. 44 consults to medical genetics were performed, and 9 to
General surgery. Clinic activity generated 101 surgeries including 37
colectomies and 7 duodenectomies. If all the appointments were done
separately this would mean at least 967 separate visits.
Conclusion The High Risk Clinic is a valuable resource for patients,
insurers and registry workers.
APC germline allele-specific expression in familial
adenomatous polyposis
E. Castellsague1, S. Gonzalez1, I. Blanco2, E. Guino3, C. Lazaro1,
S. Gruber4, G. Capella1
1Laboratori de Recerca Translacional, Institut Catala d’Oncologia,
IDIBELL, Hospitalet de Llobregat, Spain; 2Unitat de Consell Genetic,
Institut Catala d’Oncologia, IDIBELL, Hospital Duran i Reynals,
Hospitalet de Llobregat, Spain; 3Unitat de Bioestadıstica
i Bioinformatica, Institut Catala d’Oncologia, IDIBELL, Hospitalet
de Llobregat, Spain; 4University of Michigan, Ann Arbor,
MI, United States
Background About 13% of Familial Adenomatous Polyposis (FAP)
families and 70% of Attenuated FAP (AFAP) families remain with
unknown molecular pathogenic cause after APC and MYH mutational
analyses. Also, mutations can affect allele expression at the germline
level.
Aim The aim of the study was to determine the presence of germ-
line allele-specific expression (ASE) in the APC gene in FAP and
AFAP with and without detectable APC or MYH mutations.
Methods Germline RNA from fresh frozen and/or cultured lym-
phocytes of 17 APC/MYH-negative Polyposis (7 FAP, 10 AFAP)
families (21 individuals) and 26 APC-mutated Polyposis (21 FAP, 5
AFAP) families (45 individuals) was analysed. Fourteen controls
were also studied. ASE was investigated by single nucleotide primer
extension (SNuPE) of rs2229992 APC coding SNP.
Results In controls ASE was 1.05 ± 0.05. We found that 17% (3 of
17) APC/MYH(-) FAP/AFAP families showed ASE (range = 1.17–
1.39) and ASE co-segregated with disease. ASE was more intense in
short-cultured lymphocytes and reversed by puromycin treatment.
Experimental approaches are being performed to elucidate the
molecular mechanism leading to APC ASE in these families. Eleven
of 26 (42%) APC-mutated FAP/AFAP harboured ASE (range = 1.20–
8.54) being the mutant allele the under expressed one. ASE was
restricted to splicing, nonsense and frameshift mutations outside exon
15. Again, puromycin reversed ASE in all cases analysed.
Conclusions APC ASE is present in a significant proportion (17%) of
APC/MYH(-) FAP/AFAP. ASE, due to nonsense-mediated decay
(NMD), is present in APC-mutated Polyposis and is associated with
specific mutation location, similar to reports for other hereditary
syndromes. Our results show that expression-based molecular tests
could have an important role in the molecular diagnostics of Polyposis.
Is familial adenomatous polyposis a ciliopathy?
Encarna B. Gomez Garcia, Nine V. A. M. Knoers
Department of Genetics and Cell Biology, Maastricht,
The Netherlands
Familial adenomatous polyposis (FAP) is an autosomal dominant
form of intestinal polyposis and colorectal cancer caused by germ-line
mutations in the APC gene. The term Gardner’s syndrome is used to
describe a subset of families with FAP in which the extra-colonic
manifestations, such as osteomas, skin cysts, congenital hypertrophy
of the retinal pigmented epithelium (CHRPE), and desmoid tumors
(aggressive fibromatosis) are specially prominent.
Our hypothesis is that a ciliary dysfunction is the underlying
pathogenetic mechanism of the extra-intestinal manifestations
observed in patients with FAP. This hypothesis is based on the
presence of common clinical manifestations (cysts, retinal abnor-
malities, fibrosis) in Gardner’s syndrome and cilia-related disorders.
Secondly, both APC and the cilia have degradation of b-catenin as
common downstream target in the WNT-signalling pathway. Muta-
tions in the APC gene causing Gardner’s syndrome are clustered in a
region encoding a series of amino acid repeats responsible for the
binding to b-catenin. Proofs of principle that b-catenin can be the key
mediator of the ciliary disorder rely also in the observations that
overexpression of b-catenin induces polycystic kidney disease, as
well as CHRPE phenotypes in animal models. Other candidates to be
the common link are the APC-binding proteins: EB1 and Kif3a, both
of them are ciliary proteins involved in intraflagellar transport.
Finally, pathogenetic similarities between some ciliopathies and the
extra-intestinal tumors observed in FAP indicate that also the latter
can have a cilia defect. In conclusion, from the perspective of FAP as
a ciliary disorder, the presence of extra-colonic manifestations can
now be understood, and it may add a whole new range of therapeutic
options for those patients.
Adenomatous polyposis coli germline mosaicism
in a patient with classical
Judith Necker, Michele Attenhofer, Bruno Reichlin, Karl Heinimann
Research Group Human Genetics, Department of Biomedicine,
Basel, Switzerland
Familial adenomatous polyposis (FAP) is an autosomal dominant
cancer predisposition syndrome characterized by multiple colorectal
adenomas, which, unless removed, inevitably develop into colorectal
cancer. FAP is caused by germline mutations in the adenomatous
polyposis coli (APC) gene. Somatic mosaicism has been reported to
account for 11–20% of patients with apparently de novo APC
mutation. Here, we present the rare instance of a FAP patient with
APC germline mosaicism.
The index patient was diagnosed of classical polyposis and rectal
cancer at age 41. While his parents were reportedly healthy, his two
daughters displayed profuse polyposis at age 18 and 23, respectively.
Analysis of leukocyte-derived DNA from the index patient using the
protein truncation test (PTT) revealed a barely visible additional band
at 1.3 kb, indicative of a truncating mutation in the region of exon
15c–15e. By conventional sequencing of the entire coding region and
gene dosage analysis, however, no pathogenic alteration could be
identified. Subsequent APC mutation analysis of one of the patient’s
daughters revealed a strong additional PTT band at 1.3 kb resulting
from a frameshift mutation in APC exon 15c (c.2802_2805delTTAC,
p.Thr934ThrfsX19).
Abstracts 741
123
Detailed mutation analysis of DNA from epithelial and smooth
muscle portions of normal mucosa and adenomas confirmed the presence
of the c.2802_2805delTTAC alteration in the index patient. Preliminary
data from quantification analyses indicate that the proportion of mutated
alleles in these tissues varies between 13 and 46%; further investigations
on tissues originating from all three germ layers are under way.
In conclusion, our data on a FAP patient with APC germline
mosaicism
(a) highlight the importance of applying several mutation detection
methods instead of relying solely on DNA sequencing
(b) emphasize the need for early medical surveillance of the
offspring, since disease severity may increase in the next
generation, as strikingly exemplified in this family.
c.891 + 3A [ C is an Italian recurrent MutYH
mutation associated with production of aberrant mRNA
transcripts
Viel Alessandra, Pin Elisa, Pastrello Chiara, Fornasarig Mara,
Urso Emanuele, Tricarico Rossella, Tibiletti Maria Grazia,
Genuardi Maurizio
Quaia Michele Centro di Riferimento Oncologico, Aviano, Italy
In addition to p.Tyr165Cys and p.Gly382Asp, specific mutations of
the MutYH gene have been identified in different populations and
diagnostic screening strategies could be optimized accordingly. In our
mutational screening of patients with suspected MutYH-Associated
Polyposis (MAP) living in the North East of Italy, we have also
frequently identified the c.891+3A[C mutation in both homozygous
and compound heterozygous conditions. This mutation was in fact
detected 12 times in 11 probands/families and it represents the 19% of
mutated alleles in our population. The clinical phenotype of the only
homozygous c.891+3A[C/c.891+3A[C carrier, characterized by
development of less than 30 adenomas at age 51 years, was consistent
with the diagnosis of attenuated polyposis.
By RNA analysis, we demonstrated that this variant, changing the
third base of intron 10, completely disrupts the normal splicing pro-
cess, producing out-of-frame transcripts lacking exon 10 (145 bp) and
transcripts lacking exon 10 and retaining intron 11 (173 bp).
Lymphoblastoid cell lines from one homozygous and 4 compound
heterozygous patients have been obtained and analyzed at the protein
level. Western blot analyses with a mcAb directed toward the a.a.
436–536 indicated that production of the full length MutYH protein is
completely abrogated. Loss of the C-terminal domain gives rise to a
defective protein probably unable to recognize 8-oxoG and bind
DNA, thus resulting in severely impaired DNA repair.
Despite frequent recurrence in our series of MAP patients, the
c.891+3A[C mutation has been reported only rarely in other popula-
tions. A founder effect is suspected, since all patients carrying this splice
variant live and/or originate from the same geographic area (North East
of Italy). Allelotype analyses are in progress to verify this hypothesis.
Are there age and sex effects on the influence
of MUTYH variants on colorectal cancer risk?
S. M. Farrington, E. Theodoratou, H. Campbell, A. Tenesa,
M. G. Dunlop and The MUTYH Meta-Analysis Collaboration
University of Edinburgh, Colon Cancer Genetics Group, 4th Floor
MRC Human Genetics Unit, Edinburgh, Scotland
Studies into the function of the Base Excision Repair pathway and its
role in oxidative damage repair have been fuelled by the identification
of MUTYH defects predisposing to colorectal neoplasia. Initial
studies identified that bi-allelic germline MUTYH mutations were
responsible for a proportion of attenuated FAP families, which led to
the term MUTYH-associated polyposis (MAP) syndrome. However,
colorectal cancer case–control studies have demonstrated strong
evidence that bi-allelic defects of MUTYH also predispose to cancer
in the absence of multiple polyps. The question remains as to whether
mono-allelic defects impart an increased risk to disease. Various
studies have provided contradictory evidence to the role of mono-
allelic defects but the rarity of the 2 most common mutations (Y176C
MAF * 0.005; G393D MAF * 0.01) reduces the power to detect
weaker effects. Hence in order to refine the risk of bi-allelic defects
and understand the role of mono-allelic defects, in December of 2006
we invited world-wide groups, which had published data on MUTYH
analysis of colorectal cancer cases and controls, to participate in a
large meta-analysis of their data sets. We were also approached by a
number of other groups who were in the process of screening for
MUTYH defects giving a total of 10 participating groups. We are
currently in the final stages of analysis and we would like to present
our findings on the role of bi-allelic and mono-allelic MUTYH
mutations on colorectal cancer risk after analysis of the whole dataset
(consisting of 16482 cases and 14732 controls) and by age and sex
stratification.
Allele-specific expression of the APC gene in mutation-
negative patients with adenomatous polyposis
Stefanie Vogt, Astrid Kaufmann, Dietlinde Stienen, Nils Rahner,
Verena Steinke, Markus M. Nothen, Per Hoffmann, Stefan Aretz
Institute of Human Genetics, Bonn, Germany
Background In a substantial number of patients with colorectal
adenomatous polyposis the genetic basis of the disease is unknown.
Recent studies indicate that reduced expression of tumour suppressor
genes may have pathogenic relevance.
Methods We examined the allele-specific expression of the APC
gene in unselected patients with [10 synchronous colorectal adeno-
mas, in whom no APC or MUTYH germline mutation was identified,
by use of a SNaPshot analysis, a primer extension method. In 31
unrelated patients and 10 normal controls who were informative for
two APC polymorphisms the ratio of the peak areas of the different
alleles (nucleotide ‘‘C’’/‘‘T’’ in codon 486 and ‘‘G’’/‘‘A’’ in codon
535) in cDNA relative to genomic DNA (gDNA) extracted from
blood samples was determined.
Results In controls, the median cDNA/gDNA peak ratio was 0.89
(SD ± 0.13). 8/31 patients (26%) showed reduced (peak ratio
≤0.6) allelic mRNA expression, the degree of reduction ran-
ged from 40 to 72%. In 3 patients results were inconsistent; the
remaining cases had balanced expression. 6/8 cases showing reduced
expression had an attenuated, 2 had a classical colorectal phenotype.
Conclusions These findings suggest that reduced mRNA expression
may be causative for the development of a polyposis in a subset of
mutation-negative patients. Unbalanced allelic mRNA expression
point to cryptic germline mutations in the APC gene or associated
regulatory regions that are not detectable by standard methods of
mutation analysis. However, the underlying mechanisms remain to be
uncovered, yet.
The study was supported by the Deutsche Krebshilfe (Grant no.
108421)
742 Abstracts
123
Association between colorectal cancer and MYH
mutations in sporadic tumors and in attenuated familial
polyposis coli (AFAP)
E. Urso1, M. Agostini1, S. Pucciarelli1, I. Mammi1, A. Viel2,
I. Maretto1, D. Nitti1
lClinica Chirurgica 2^, Dipartimento di Scienze Oncologiche
e Chirurgiche, Azienda Ospedaliera-Universita di Padova, Italy;2Oncologia Sperimentale 1^, Centro Regionale Oncologico, IRCCS,
Aviano, Italy
Background While MYH biallelic mutations are clearly related to
the Attenuated Familial Adenomatous Polyposis (AFAP), the asso-
ciation to an increase risk of colorectal cancer (CRC), and the
phenotype of monoallelic MYH mutations are still unclear.
Aim To Investigate in a prospective series of CRC and AFAP
patients the association between MYH mutations and colonic tumors.
Methods Study population consisted in a consecutive unselected
series of CRC patients who underwent surgery in 2003 and a consec-
utive cohort of 37 AFAP patients observed from 2003 to 2008 at Padova
Hospital. All patients were screened for the most frequent MYHgermline mutations (Y165C, G382D, IVS10+3A[C, 1395-7delGGA).
Results Of 430 CRC patients [F = 170; median age 67 years], 33
were B50 years old (yo). Two biallelic and 2 monoallelic MYH
mutations were found and 1 patient B50 yo had monoallelicmutation
(3% of the early onset CRC). 37 AFAP patients [F = 11; median age
47 years] were observed; median (range) polyps number was 40
(10–100), and 3 APC mutations were found. Seventeen AFAP
patients had CRC and 11 of them were B50 yo. MYH biallelic and
monoallelic mutations were found in 6 and 3 patients of the whole
series, respectively. The corresponding figure for AFAP patients with
CRC were 3 and 2, respectively; and the corresponding figure for
AFAP patients with CRC B 50 yo were 2 and 1, respectively. Inci-
dence of MYH biallelic and monoallelic mutation is not statistically
different in AFAP patient withor without cancer, even considering
AFAP patients with CRC B 50 yo.
Conclusions The Incidence of MYH mutations is low in unselected
CRCs patients; however it is not so negligible in patients B50 years.
In AFAP patients, the presence MYH mutation (either mono or
biallelic) is not associated with an increase incidence of CRC, also
considering patients B50 yo.
Changing trends in causes of mortality for patients
with familial adenomatous polyposis
Ashish Sinha, S. Rashis, R. K. S. Phillips, S. K. Clark
St Mark’s Hospital, Polyposis Registry, St Mark’s Hospital,
Harrow, London, UK
Introduction Widespread use of prophylactic colectomy in familial
adenomatous polyposis (FAP) has resulted in reduction in the number
of deaths due to colorectal cancer. Now extracolonic manifestations of
FAP are thought to be the leading cause of mortality in these patients.
Methods We retrospectively examined the cause of death in patients
with a confirmed diagnosis of FAP who had undergone primary
surgery at our institution, using the Polyposis Registry database.
Results Of the 559 patients who underwent primary surgery for FAP
at our institution 146 (70 male) have since died. Information was
available on 143 patients. The median age of death before 1990 was
45 years (interquartile range 35–54) and since 1990 was 52 years
(40–62); p = 0.022 (Mann–Whitney U-test). Metastatic colorectal
cancer caused 53 (37.1%) deaths at a median age of 43 years (35–54).
Eleven of these patients had known cancers at the time of surgery, 42
subsequently developed rectal cancer and metastatic disease. Desmoids
caused mortality in 16 (11.2%) at a median age of 34 years (25–42).
Upper gastrointestinal malignancies caused 26 (18.2%) deaths at a
median age of 53 years (43–66). Other causes of mortality included 28
(19.6%) non-FAP related deaths at a median age of 54 years (45–66) and
7 (4.9%) secondary to non-FAP related malignancies; median age
52 years (49–60). There were 13 (9.1%) perioperative mortalities. Nine
(11.5%) and 4 (6.2%), p = 0.03 (Mann–Whitney U-test) were pre and
post 1990 respectively. These occurred at a median age of 34 (25–39)
and 53 (42–55), p = 0.01 (Mann–Whitney U-test) pre and post 1990
respectively.
Conclusion Metastatic colorectal cancer remains the leading cause
of death in FAP. In recent decades more deaths are attributable to
extracolonic manifestations. Cardio-respiratory disease is the leading
cause of non-FAP related death. Post 1990, surgery appears to be
safer with significantly fewer perioperative mortalities.
How a nurse practitioner specialising in pollyposis
supports both patients and doctors
Muditha Samarasinghe, Kay Neale, Pam Nye
The Polyposis Registry, St Marks Hospital, Harrow, Middlesex, UK
Medical staff come to our hospital to gain experience in colorectal
conditions. They stay for a maximum of 1 year but often only
6 months. On arrival they have rarely encountered patients with
polyposis and know little of these syndromes. When they leave, they
take their valuable, newly acquired knowledge with them. The
patients inevitably suffer from this fluctuating level of experience.
In her clinic the Nurse Practitioner sees patients who are attending
for routine follow up. She carries out a full physical examination,
reviews the endoscopy and other investigative reports. The clinics run
concurrently with the Consultants’ clinics in order to provide
immediate expert advice if patients present with new findings. On
average 500 patients a year will be seen by the Nurse Practitioner
alone. In addition medical staff will request a joint consultation with
the Nurse for a similar number. Advice is given on things such as:
• Screening intervals
• Familial tendencies
• Genetic variation
• Genetic testing
• Screening protocols for desmoid disease or adrenal adenomas
The Nurse will also take over from the doctor to provide psy-
chological support when necessary releasing the doctor for the next
patient.
Between clinic visits these patients require easy access to spe-
cialist medical advice. In addition they have particular psychological,
emotional and social needs requiring sensitive management. The
Nurse Practitioner works closely with the Nurse Specialist and
Administrative staff and:
• a telephone and email helpline are provided
• patients who miss an appointment are contacted; the importance
of screening is discussed and another appointment offered
• customised clinic lists are produced to alert the nurses to specific
points to be covered at individual patients’ visits
Conclusion Polyposis syndromes are complex; patients’ lives are
enhanced by continuity of care and doctors are helped by a specially
trained nurse.
Abstracts 743
123
‘‘Peyer’s patch’’: Two cases in familial adenomatous
polyposis patients after colectomy
R. Man
St. Mark’s Hospital, Harrow, Middlesex, UK
Introduction Peyer’s Patches, gut-associated lymphoid tissue
(GALT), are lymphoid follicles (Gullberg & Soderholm 2006). They
are associated with Crohn’s disease (Shikuwa et al. 2007), malignant
lymphoma (Rappaport et al. 1971), idiopathic intussusception
(Hasegawa et al. 1998) and spongiform encephalopathy. They often
described as polyps and may be mistaken for dysplastic lesions.
Aim and method This study aimed to report and describes the mac-
roscopic appearance of Peyer’s patch using various endoscopic imaging
techniques in two FAP patients with ileoanal pouch. A retrospective
search of our endoscopy database identified two FAP patients (26 years
old and 46 years old respectively) both with an atypical ileal lesion
found during surveillance endoscopy (15 years and 2 years after pouch
surgery respectively). The macroscopic appearance and characteriza-
tion of these lesions was examined with three different imaging
techniques: conventional endoscopy with white light, chromoendos-
copy with indigo carmine and electronic chromoendoscopy with narrow
band imaging. Biopsy was taken to for histological assessment.
Results Both lesions appeared to be elevated from the surface
mucosa. Round lymphoid follicles were identified on close observa-
tion. Enhanced imaging with indigo carmine and narrow band
imaging ascertain the absence of dysplastic features. No excision
applied to any of these lesions. Histology later confirmed the present
of lymphoid follicles only.
Conclusion Recognition of different lesions is one of the core
objectives in endoscopy training. Advances in endoscopy, such as
video capsule endoscope and double balloon enteroscopy, increase
the observation of these unusual lesions in the more accessible distal
ileum; in particular in FAP after colectomy with easy access to the
distal ileum. Unusual lesions may not be familiar to some endosco-
pists. Descriptions of macroscopic appearance of Peyer’s patch in
endoscopy are limited in the literature (Hizawa et al. 1996, Fujimura
et al. 1996, Fujimura & Owen 2000). Accurate and early identification
and differentiation of the types of pathology may allow prompt
treatment and management. Inability to recognize these lesions may
lead to unnecessary excision and the associated risks and complica-
tions. Enhanced imaging techniques may allow detailed interpretation
of mucosal lesions, and may help reducing this problem. Peyer’s
patch can be found in normal healthy person. Implication of Peyer’s
Patch in FAP patient is not clear.
Completion proctectomy: three case studies
in FAP patients with ileo-rectal anastomosis
R. Man, K. Neale, S. K. Clark
St. Mark’s Hospital, Harrow, Middlesex, UK
Introduction FAP patients with ileo-rectal anastomosis need regular
surveillance of their retained rectum. Any detection of high risk
dysplastic lesions requires prompt excision. The choice of manage-
ment could vary individually and sometimes could be difficult. This
study attempted to highlight the management of three cases with high
risk rectal lesions.
Aim and method This study aimed to explore the choice of man-
agement in patients with high risk rectal lesions detected during
endoscopic surveillance. A retrospective search from the St. Mark’s
polyposis registry identified three cases. Their management choices
and its implications were examined.
Case 1 A 55 year old man with FAP had a rectal lesion removed
during a surveillance endoscopy, 29 years after IRA. Histology con-
firmed invasive carcinoma. Screening imaging including endorectal
ultrasound, CT scan and blood tests found no evidence of local
invasion or metastasis after endoscopic polypectomy. Hence, proc-
tectomy was not performed. The patient died 10 years later from a
malignant kidney tumour.
Case 2 A 65 year old women with FAP was found to have a
severely dysplastic lesions in the rectum, 22 years after IRA. The
rectal lesion was removed endoscopically. Completion proctectomy
was declined by patient. No further high risk rectal lesion was
detected with an endoscopic follow up of 4 years.
Case 3 A 62 year old women with FAP had multiple severely
dysplastic rectal lesions detected 2 years after the construction of
IRA. Proctectomy was attempted but failed due to the presence of
extensive desmoid disease. Learning difficulties also limited her
choice for end-ileostomy. She settled with regular endoscopy and the
use of chemoprevention as a result. Seven years later, repeated his-
tological reviews of the lesion remain severely dysplastic.
Conclusion Completion proctectomy is usually suggested when
rectal cancer is detected in FAP patients after ileo-rectal anastomosis.
Advances in therapeutic endoscopy allow safe and effective excision
of mucosal lesions including polyp cancer. It is not unknown whether
surgery can be delayed if the rectal lesion can be completely removed
locally. High risk rectal lesion in IRA is the most common predicting
factor and indication for surgery. Not all IRA patients with high risk
rectal lesion proceed to proctectomy. When deciding the need for
proctectomy in FAP patients with ileo-rectal anastomosis, the situation
can be complicated especially in mentally and physically compro-
mised patients. The long term risk in patients who choose not to or
failed to convert from IRA to pouch or end-ileostomy is not known.
Endoscopic monitoring and chemoprevention may be employed as
comprising options. These should be considered with caution.
Peutz–Jeghers syndrome: screening and follow-up
Shirley V. Hodgson, Andrew D. Beggs
St Georges, University of London, London, UK
Peutz Jeghers syndrome (PJS) is a rare autosomal dominant condition
characterised by the development of multiple hamartomatous polyps
throughout the bowel, and mucocutaneous pigmentation which is
variable, develops during childhood and may fade in later adulthood.
These polyps develop in early childhood, predominantly in the small
intestine but also in the large bowel and stomach.
Diagnosis is often made by obstruction from intussusception
(43%) or obstruction (23%); abdominal pain from infarction (14%) or
rectal bleeding from ulceration (7%). A recent review by Giardello
et al. estimated lifetime risk of GI cancer as 48% for the small bowel,
24% gastric, 24% colon, 5% pancreas (up to 100-fold increase in risk
reported) and female breast cancer 32% by 60 years age. There was
also an increased risk of sex cord tumours of the ovary or testis, which
can secrete estrogenic hormones, which may explain the increased
risk of adenoma malignum of the cervix. The total risk of death from
cancer in PJS has been estimated to be 48% by age 57 years.
Mutations in the STK11 serine/threonine kinase gene on chromo-
some 19p13.3 are responsible for PJS, with no clear evidence for
genetic heterogeneity, and only a minor suggestion of genotype/
phenotype correlations. Clinical management has been developed to
detect early neoplastic lesions, and includes annual clinical evaluation,
744 Abstracts
123
abdominal and pelvic ultrasound, cervical smears and breast exami-
nation, biennial ‘‘top and tail’’ endoscopy & small bowel series, and
mammography for affected women from 25 years age. Surveillance
for pancreatic cancer remains problematic, and screening protocols are
being revised in the light of clinical evidence.
We have set out to obtain follow-up data on patients with PJS
ascertained from centres around Europe to try and gain further insight
into the incidence of cancer and the effectiveness of surveillance in
affected individuals, and preliminary data from this will be presented.
The mutational spectrum of APC in Greek FAP
patients including a distinct mutation on the alternative
splice site of exon 9
Florentia Fostira1, Georgia Thodi1, George Fountzilas2,3,
Drakoulis Yannoukakos1
1Molecular Diagnostics Laboratory, I/R-RP, National Center
for Scientific Research ‘‘Demokritos’’, Athens, Greece; 2Department
of Medical Oncology, Aristotle University of Thessaloniki,
Papageorgiou Hospital, Thessaloniki, Greece; 3Hellenic Cooperative
Oncology Group, Athens, Greece
Familial Adenomatous Polyposis (FAP), an autosomal dominant
inherited disease caused by germline mutations within the APC gene,
is characterized by early onset colorectal cancer as a consequence of
the intrinsic phenotypic feature of multiple colorectal adenomatic
polyps. Analysis of the APC gene performed in a Greek kindred
consisting of twenty-five FAP families, revealed eighteen different
germline mutations in twenty (80%) families, of which five novel.
Twelve mutations are located within exon 15, carriers of which are
characterized by a clearly earlier mean age of colorectal polyposis
(30.4 years) compared to the rest, which are scattered between exons
3 and 11 (39.4 years). Interestingly, a novel mutation located on the
alternatively splice site of exon 9 was identified in a patient with
attenuated FAP phenotypic features. This is the first reported mutation
in the specific site. Transcripts characterization revealed disruption of
splicing occurring within exon 9, resulting in the expression of a
shorter mRNA transcript, which surprisingly does not affect the ratio
between the two wild type transcripts. The latter, along with the ‘three
hit theory’, highlights the need for a third hit for cancer to develop,
most probable contributing to the mildness of disease. This is the first
full report on the clinical characterization and the mutation spectrum
in Greek adenomatous polyposis families. Using all available pub-
lished data on Greek FAP families, it is concluded that the population
is characterized by genetic heterogeneity, lack of founder mutation in
FAP syndrome and low incidence of genomic rearrangements in
APC.
Your insight membership
Kay Neale, Tina Isherwood
The Polyposis Registry, St Mark’s Hospital, Harrow, Middlesex, UK
In Japan in 2007 we presented a Poster to tell people about InSiGHT.
It was felt important that current members should be helped to
understand how the Society is organised and why they should con-
tinue to pay an annual subscription. Also, non-members of the Society
attending the Scientific Meeting would be encouraged to join and told
how to apply for membership.
It is proposed that the poster giving the details listed below should
be updated and presented again in Germany in 2009:
1. Names of the members of Council
2. The way in which the Officers and members of Council are
elected
3. How Council is funded
4. Number of current members by country
5. Subscription rates and methods of payment
6. Distribution of the Journal
7. The database
8. Website address
9. Request for suggestions
Bowel phenomena and symptom experience
after 6 months of total colectomy with familial
adenomatous polyposis
Yoshie Murakami
Dult Nursing, Graduate School of Human Health Sciences,
Tokyo Metropolitan University, Tokyo, Japan
Purpose The purpose of this study was to explore FAP patients’
bowel phenomena and symptom experience during the first 6 months
after undergoing a prophylactic colectomy.
Methods A convenience sample of three patients, one female and
two males were interviewed, using a semi-structured interview guide,
based on the UCSF Symptom Management Model.
Results The data revealed that three patients had six common bowel
phenomena; defecation patterns, sense for defecation, ability to dis-
criminate between stool and flatus, soiling, awareness of abdominal
content movement (peristalsis), and awareness of bowel sounds. The
data also revealed that each patient had unique perception of their
symptom experience; diarrhea, fear of bowel blockage, and night
soiling. All patients had cope with symptom experience and thought
value of life, because they could defect colon cancer and not enough
to death.
Conclusion Nurses, to provide quality nursing care to FAP patient
post-colectomy, need to become aware of each FAP patient’s unique
perception of their symptom experience, as well as the bowel phe-
nomena common to all such FAP patients.
MUTYH variants in Austrian patients with familial
adenomatous polyposis-like symptoms: a first report
Brigitte Wolf, Silke Gruber, Judith Karner-Hanusch
Medical University of Vienna, Department of Surgery, Research
Laboratories, Wien, Austria
Background In the diagnosis of Familial Adenomatous Polyposis
(FAP) a number of patients present an atypical medical history
characterized by less polyps and an older age of onset. Some of these
patients have been shown to bear an inherited defect in the base
excision repair gene MutYH.
Materials and methods We analyzed a series of 64 unrelated Aus-
trian families clinically diagnosed with FAP (17; 26.6%), AFAP (31;
48.4%) or multiple colorectal adenomas (16; 25%) for mutations in
the MutYH gene. All 16 exons of the gene were sequenced starting
from genomic DNA. An inherited defect in the APC gene was
excluded by sequence analysis and MLPA.
Results and conclusion In 15 of 64 (23.5%) patients a mutation in
the MutYH gene was identified. In nine (14.1%) patients both alleles
Abstracts 745
123
were affected. Among the mutation carriers three (20%) patients
presented classical FAP symptoms, 10 (66.7%) patients AFAP and
two (13.3%) multiple colorectal adenomas. Three of the mutation
carriers had polyps in the upper intestine and three developed colo-
rectal cancer before the age of 50 years. Only one of the putative
pathogenic missense variants has not been listed in the MutYH
database before, but was detected in both alleles of an AFAP patient.
Furthermore we identified 5 frequently reported variants and one
variant 127 bp upstream of the gene.
This study is the first comprehensive report of MutYH gene
mutations in Austrian FAP and FAP-like patients.
The Lithuanian polyposis register: progress during
past 14 years
N. E. Samalavicius, T. Poskus
Oncology Institute of Vilnius University and Vilnius University
Santariskiu Clinic Center Branch, Vilnius, Lithuania
Introduction The Lithuanian Polyposis Register has been found in
1995. Up till then, no attempts for systematic registration, screening,
treatment or follow-up have been made. The support for the initiation
of the register has been received from EUROFAP project, and register
itself has been established in a close collaboration with the Leeds
Castle Polyposis Group.
Materials and methods During the period January 1995 to Decem-
ber 2008, a total of 106 familial adenomatous polyposis (FAP)
patients from 47 unrelated families were registered. Among them, 22
(46.8%) were isolated cases. Data on screening, surgical treatment,
extracolonic manifestations and outcome have been investigated.
Results Out of 106 registered FAP patients, 29 (27.4%) underwent
prophylactic surgery, 49 (46.2%) were operated in the presence of
colorectal cancer, and 18 (26.4%) were not operated. Methods of pro-
phylactic treatment included either subtotal colectomy with caecorectal/
ileorectal anastomosis, or reconstructive proctocolectomy. Most of the
patients with colorectal cancer underwent segmental resections.
FAP patients were examined for extracolonic manifestations: in
94.7% of CHRPE were detected, in 68.2%—mandibular osteomas, in
only 14.3%—fundicgland polyposis, and in 66.7% duodenal adeno-
mas. Desmoid tumours were present in 7.1% of FAP patients, and
14.3% had epidermoid cysts. No cases of duodenal cancer or other
site cancers were recorded.
Conclusions The Lithuanian polyposis register has been success-
fully growing since 1995. In a country like Lithuania with
approximately 3 million inhabitants, a centralized register has been
proven to be the optimal solution. However, proper molecular genetic
services are yet to be established.
The role of the nurse specialist at St Mark’s Hospital
Jo Rawlings, Kay Neale
The Polyposis Registry, St Mark’s Hospital, London, UK
The role is much the same as in Registries the world over but in this
job the nurse attends surgical clinics where patients with polyposis are
also being seen. There are six clinics a week and a paediatric clinic
once a month. In the last year 1,165 patients attended, 497 of these
were seen by the nurse, 115 of whom had genetic counselling. In
addition to this and the routine of taking a family history, there is an
opportunity to meet families over and over again at follow up visits,
or when they attend with relatives, to get to know them well and earn
their trust. Conversations starting about football, pets or music
sometimes lead to things rarely discussed or forgotten such as ille-
gitimacy, infantile death or relatives distanced by family feud.
Another advantage of working within the hospital is the oppor-
tunity to visit patients in the ward where they have time to talk. This is
a good time to update the family pedigree, as relatives with a more
extended knowledge may be visiting, and also to re-enforce infor-
mation given previously at clinic visits.
In addition to face to face contact time is spent in the office where
advice and support can be given by telephone and e-mail and efforts
are made to trace at risk relatives. Information obtained in the clinical
setting is entered onto the Registry database and family trees drawn.
A poster would give the opportunity to present more information
about the role, including a case history of a young foreign national
with a fondness for vodka who presented in A&E with pancreatitis
and anaemia. Fortunately, he was seen by a knowledgeable doctor and
FAP was diagnosed. The nurse was instrumental in ensuring that this
extremely anxious and reluctant young man attended for surgery and
subsequent care.
Developing a protocol for genetic evaluation
of APC-negative patients with multiple colorectal
adenomas
Guy Rosner1, Dani Bercovich2, Hana Strul1, Revital Kariv,
Zamir Halpern1, Paul Rozen1
1Department of Gastroenterology, Tel Aviv Medical Center,
Tel Aviv, Israel; 2Human Molecular Genetics & Pharmacogenetics,
Migal, Galilee Bio-Technology Center
Background Genetic evaluation of APC-negative patients with
multiple colorectal adenomas (MAP) is a clinical & economic burden.
In Israel, this is confounded as HMO-financed APC genetic evalua-
tion sequences exon 16 only up to 3,000 base-pairs from 50.
Aims Perform comprehensive genetic evaluation of APC-negative
MAP patients & draw conclusions for a future evaluation protocol.
Methods Study population: 29 Amsterdam-criteria negative Jewish
individuals, 52% males of mean age 53 years (range 19–76). There was
dominant neoplasia inheritance in 7, recessive in 14; 12 reported CRC in
a first-degree relative. Numbers of adenomas were: 6–10 (3 patients, all
with colorectal cancer [CRC]), 11–19 (7 patients), multiple (19 cases); 7
had CRC (6 in left colon). Genetic analyses included: completion
of exon 16 sequencing, MLPA analysis for deletions/duplications,
MUTYH gene sequencing, MSI in CRCs having Bethesda criteria.
Results Exon 16 sequencing revealed a weakly pathogenic poly-
morphism. MLPA results are pending. Pathogenic MUTYH mutations
were found in 3 patients; 2 others had variants of unknown significance;
polymorphisms occurred in 12, 5 having[1. 1 patient was MSI-H with
MLH1 immunohistology, 5 more Bethesda-positive CRCs await MSI�.
Origin Exon 16
sequencing
APC
MLPA
MYH
mutation
MYH
variants
MSI-H�
IH+
European 11 Normal Pending None None
N. African 9 Normal Pending 3 (33.3%) 1 (11%)
Middle-
Eastern 9
Normal Pending None 1 (11%) 1 (11%)
746 Abstracts
123
Discussion Based on this small, APC-negative, MAP cohort: Lynch
syndrome needs exclusion if MAP is associated with CRC; otherwise
MUTYH sequencing is probably the first evaluation especially for
Jews of North-African descent. The significance of MUTYH variants
needs further assessment; MUTYH polymorphisms might serve as
MAP modifiers. Sequencing exon 16 is relatively unrewarding. We
await MLPA & MSI results to draw final conclusions on a MAP
evaluation protocol. Supported by the Israel Cancer Association &
Sistopali Fund for Gastrointestinal Cancer Prevention.
Ethnic variation of MUTYH gene mutations
in an Israeli population sample
Guy Rosner1,2, Sharon Simchoni2, Avi Orr-Urtreger2, Ruth Shomrat2,
Dani Bercovich3, Hana Strul1, Revital Kariv1, Zamir Halpern1,
Paul Rozen1
1Department of Gastroenterology, Tel Aviv Sourasky Medical Center,
Tel Aviv, Israel; 2Department of Genetics, Tel Aviv Sourasky
Medical Center, Tel Aviv, Israel; 3Human Molecular Genetics
& Pharmacogenetics, Migal, Galilee Bio-Technology Center
Background MUTYH gene mutations predispose to colorectal pol-
yposis (MAP). Mutations Y179 C (exon 7) and G396D (exon 13), are
the most frequent, however novel variants occur in defined populations.
Aims Analyze our initial results evaluating MUTYH mutations in an
Israeli population.
Methods These were 65 Jewish patients of different origins, having
[5 colorectal adenomatous polyps & negative for APC mutations. All
had sequencing of MUTYH exons 7 and 13, & 29 also had full gene
sequencing.
Results 26 abnormal alleles were found, 22 known to be pathogenic.
There were 8 biallelic mutations (6 G396D homozygotes, 2 Y179C
homozygotes) & 1 compound heterozygote (G396D/Y179C); 4 mono-
allelic mutations (1 G396D, 2 Y179C, one 1186–1187 insGG). 4
others had MUTYH variants of unknown significance (L406V, L401V,
L417M, S512F). 16 polymorphisms were found in 12 patients with 5
having[1 polymorphism. 20 of 26 abnormal alleles were in Jews of
North-African origin. No Jews of European origin had either of the 2
common pathogenic mutations.
Origin/No. pts. G396DY179CNo. alleles/biallelic
Other path.mutationsNo. alleles
Variants ofunknownsignificance
Polymor-phisms No.persons/no.with [1
N. African/27 10/4 7/2 1 2 3/1
European/14 0 0 0 0 6/3
Middle East/24 4/2 0 0 2 3/1
Discussion Based on this pilot study, MUTYH gene mutations are
not uncommon in Israeli Jewish APC-negative MAP patients. Pub-
lished experience of MUTYH gene mutations in Jews refers mainly to
those in North America & usually of European-origin. In this initial
study the two most common mutations were found in Jews of non-
European origin. The significance of MUTYH variants & biallelic
polymorphisms needs further assessment in our population. Further
investigation for MUTYH mutations in Jews of various origins is
warranted.
Supported, in part, by the Sistopali Fund for Gastrointestinal Cancer
Prevention.
Is the ‘mutated in colorectal cancer’ gene important
in colon cancer after all?
L. Pangon, N. Sigglekow, E. A. Musgrove, M. Kohonen-Corish
Garvan Institute of Medical Research, Sydney, Australia
The ‘mutated in colorectal cancer’ (MCC) gene was discovered in
1991 due to its close linkage with APC, during the search for the FAP
susceptibility gene. Although MCC mutations were found in sporadic
cancers, early attempts failed to determine the significance of an
MCC defect in colon cancer.
We have recently shown that the MCC promoter is hypermethy-
lated in *50% of colon cancers and therefore this gene defect is more
common than previously thought (Kohonen-Corish et al. 2007;
Oncogene 26:4435). MCC methylation is strongly associated with the
BRAFV600E mutation, CIMP+ and MSI-H phenotypes. These find-
ings were recently confirmed in an independent patient cohort
(Fukuyama et al. 2008; Oncogene 27:6044). MCC methylation occurs
early in precancerous lesions and is more common in serrated polyps
than in traditional adenomas, suggesting a role in the serrated neo-
plasia pathway. MCC is methylated independent of the adjacent APC
gene, and therefore it is not a bystander effect.
Previous studies indicate that MCC has a potential role in regu-
lating cell cycle progression and the NFkB and WNT pathways. All
these cellular processes are relevant in carcinogenesis and defects of
each one have been described in colon cancer. We have extended
these initial observations and further dissected the effect of MCC on
the cell cycle. Our results are consistent with a previously reported
role at the G1/S checkpoint and a potential new role at the G2/M
checkpoint. Our mass spectrometry experiments identify new MCC
interacting partners that support its role in the cell cycle. We have
further refined the subcellular localisation of MCC and determined
that its activity may be regulated through phosphorylation of specific
amino acids in a cell cycle dependent manner.
Our data provide a potential cellular mechanism whereby an MCC
defect promotes colon cancer.
Mucosectomy and stapled pouch-anal anastomosis
Steffen Bulow
The Danish Polyposis Register, Copenhagen, Denmark
Background In comparison with IRA the advantage of an ileoanal
pouch is the total removal of all premalignant rectal mucosa. In FAP
most surgeons prefer to perform a mucosectomy with a hand-sewn
pouch-anal anastomosis, although the function is better after a stapled
anastomosis. We have tried to combine the advantages of a muco-
sectomy and a stapled anastomosis.
Procedure Proctocolectomy is performed including rectal transsec-
tion at the pelvic floor and construction of a stapled J-pouch with the
anvil of a circular stapler 29 mm inserted in the bottom of the pouch.
A Lone star retractor is applied and the rectal mucosa is incised
10 mm above the dentate line. Mucosectomy is performed, and a
purse-string suture is applied to the resection line. The circular stapler
is introduced transanally, connected to the anvil and the purse string
suture is ligated around the center rod. The stapler is pressed firmly
upwards, approximated, fired and removed, thereby leaving a stapled
Abstracts 747
123
anastomosis at the level of the dentate line. A protective loop ileos-
tomy is used routinely.
Results Ten patients (7 males and 3 females, median age 37 years,
range 12–50) were operated: 6 with a proctocolectomy and 4 with a
proctectomy after a previous IRA. One patient developed a small
abscess behind the pouch, which was drained successfully, and in two
patients an anastomotic stricture necessitated one and two dilatations,
respectively. Two patients have not yet had their ileostomy closed.
After a median observation of 25 months (range 3–121) 8 patients are
fully continent without any urge and have a median of 5 bowel
movements per day (range 3–8). No anastomotic adenomas have been
observed at regular endoscopical follow-up.
Conclusion Mucosectomy with a stapled pouch-anal anastomosis
seems to result in good function without any increased risk of anas-
tomotic adenoma formation in the short term.
Mutation analysis of the APC gene in Taiwanese FAP
families; low incidence of apc germline mutation in FAP
families with mixed type of polyps
Jy-Ming Chiang, P. S Hsieuh, C. R. Chang-chen
Chang Gung Memorial Hospital Tao-Yuan, Taiwan
Purpose Familial adenomatous polyposis (FAP) is an autosomal
dominant disease caused by germline mutations in the adenomatous
polyposis coli (APC) gene. Affected individuals develop colonic pol-
yposis and various extra-colonic manifestations. This study is aimed to
investigate the genetic and clinical characteristics of Taiwanese FAP
families, and to analyze genotype–phenotype correlations.
Materials and methods Blood samples were drawn from patients
diagnosed with classic FAP registered in hereditary colorectal cancer
database. Mutation analysis of APC gene of 66 FAP patients from 47
unrelated families was first screened. Negative cases were tested with
Multiplex ligation-dependent probe amplification (MLPA) and SSCP
of MYH exon 7 and 13.
Results 79% (37/47) families had 28 APC mutations including 19
frameshift mutations, 4 nonsense mutations, 3 genomic deletion
mutations, 1 missense mutation and 1 splice site mutation. We iden-
tified 15 novel mutations in 32% (15/47) families. Patients without
identified APC mutation significantly displayed less profuse Polyposis
(p = 0.034) and less gastroduodenal polyps (p = 0.027). Further-
more, FAP families with mixed types of polyposis are significantly
associated with low incidence of APC germline mutation (p = 0.002).
Conclusions We added APC germline mutation data of Taiwanese
FAP patients to the world and indicated that clinically defined FAP
family may demonstrate distinct variant that is less frequently caused
by gremlin mutation of APC gene.
Location in the large bowel influences the APC
mutations observed in FAP adenomas
O. Will, S. J. Leedham, G. Elia, R. K. S. Phillips, S. K. Clark, I. P. M.
Tomlinson
St Mark’s Hospital, Middlesex, UK
Introduction The right colon differs from the left, in embryological
origin, luminal environment, and function. In both sporadic colorectal
cancer and Familial Adenomatous Polyposis (FAP), polyp density and
cancer susceptibility vary markedly by colonic site. Adenomas in
FAP have a different mutational spectrum in small intestine versus
colon. This study aimed to investigate whether colonic location also
influences the APC mutation spectrum in FAP.
Methods 127 1–2 mm mildly dysplastic adenomas from 5 patients
with a codon 1309 germline mutation, and 41 from 3 patients with
mutations proximal to codon 1265, were analysed to assess the fre-
quency of loss of heterozygosity (LOH). We chose polyps from
different locations in the colon. Immunohistochemistry for beta-
catenin, caspase-3 and Ki-67 was performed to assess WNT pathway
activation, apoptosis and proliferation.
Results In polyps from patients with a 1309 mutation, the frequency
of LOH showed a gradient from rectum (highest) to caecum/
ascending colon (lowest), but this was not present in patients with
proximal germline APC mutations. Crypt-by-crypt analysis confirmed
the LOH findings from whole polyps. Beta-catenin and caspase-3
expression showed no significant variation by colonic region, but
Ki-67 expression decreased from ascending colon to rectum in
tumours and normal tissue.
Conclusions Colonic site alters the mutational spectrum of APC,
and crypt cell proliferation. The higher frequency of LOH in rectal
polyps from patients with codon 1309 mutations may help to explain
their increased polyp burden at this site compared with patients who
have other germline APC mutations.
748 Abstracts
123