Download Congenital hypertrophy of the retinal pigment epithelium in familial

2400
Congenital Hypertrophy of the Retinal Pigment
Epithelium in Familial Adenomatous Polyposis
Novel Criteria of Assessment and Correlations with Constitutional Adenomatous
Polyposis Coli Gene Mutations
Rosa Valanzano, M.D.'
Alessandro Cama, M.D?
Roberto Volpe, M.D?
Maria Cristina Curia, Ph.0.'
Rita Mencucci, M.D?
Raffaele Palmirotta, M.D?
Pasquale Battista, Ph.o.2
Ferdinand0 Ficari, M.D.'
Renato Mariani-Costantini,
Francesco Tonelli, M.D.'
M.D.'
' Dipartimento di Fisiopatologia Clinica, Unita'
di Chirurgia, Universita' di Firenze, Firenze, Italy.
Cattedra di Patologia Generale, lstituto di Patologia Umana e Medicina Sociale, Universita'
"Gabriele D'Annunzio", Chieti, Italy.
Clinica Oculistica I, Universita' di Firenze, Firenze, Italy.
Preliminary data on congenital hypertrophy of
retinal pigment epithelium have been published
in: L'ipertrofia congenita dell'epitelio pigmentato della retina quale sensibile marker della
poliposi familiare del colon. Chirurgia
1989: 2(9):472-6. Mutations of the adenomatous polyposis coli gene have been published
as precised in the text and references.
This work was supported by A.I.R.C. special
project "I tumori ereditari del colon," by CNRACRO Grant 94.01 161.PF39, and by M.U.R.S.T.
40%.
Address for reprints: Rosa Valanzano, M.D., Dipartimento di Fisiopatologia Clinica, Unita' di
Chirurgia, Viale Morgagni, 85. 50134, Firenze,
Italy.
Received May 3,1996; revision received August
12, 1996; accepted August 12, 1996.
0 1996 American Cancer Society
BACKGROUND. Congenital hypertrophy of the retinal pigment epithelium (CHRPE)
is the most common extracolonic manifestation of familial adenomatous polyposis
(FAP) and is an early clinical marker of the disease. It seems to be correlated with
the position of constitutional mutations of the adenomatous polyposis coli (APC)
gene.
METHODS. The authors investigated the expression of CHRPE and its correlation
with the position of the APC gene in FAP patients and in "at risk" relatives from
31 FAP kindreds. To obtain comparable data on CHRPE expression, the authors
developed a novel scoring system based on morphologic and dimensional criteria.
RESULTS. A positive CHRPE score was obtained in 29 of 39 FAP patients (74%) and
in 16 of 53 relatives who showed no clinical evidence of FAP (30%). Colonoscopy
revealed polyps in 20 of the 47 relatives who could be examined. The cumulative
sensitivity and specificity of CHRPE were 72.88% and 96.29%, respectively. APC
gene mutations were characterized in 34 subjects from 17 kindreds. In 28 of the
subjects, mutations were detected in exon 15, between codons 876 and 1324.
Mutations were found in exon 9 in 6 subjects. In 3 of the 6 subjects, they were
found at the site where both forms of alternative splicing of the exon occur (codon
437). In the other 3 subjects (another kindred), mutations were found in the portion
of exon 9 in which alternative splicing occurs (codon 367). Only 1 of the 6 subjects
(16.6%) with mutations in exon 9 had a positive CHRPE score, compared with 28
of 28 subjects (100%) with mutations in exon 15. None of the 3 subjects with
mutations in codon 437 had a positive CHRPE score. The CHRPE scores of exon
15 mutation carriers varied markedly both within and among kindreds, irrespective
of the mutation site.
CONCLUSIONS. The results of this study indicate that the site of APC gene mutation
influences CHRPE expression but is not the only factor responsible for the presence
and level of retinal lesions in FAP patients. Cancer 1996; 78:2400-10.
0 1996 American Cancer Society
KEYWORDS: polyposis, familial adenomatous, hypertrophy, retinal pigment epithelium, congenital, adenomatous polyposis coli gene, mutations.
F
amilial adenomatous polyposis (FAP) is a dominant inherited disease
characterized by the development of multiple colorectal adenomas
that usually arises in the second decade of life. However, age at disease
onset and the number and distribution of polyps vary among patients,
even within the same family.'-3 Phenotypic variability is also evident for
the extracolonic lesions associated with FAF', which include epidermoid
cysts, osteomas, dental abnormalities, desmoid tumors, and congenital
hypertrophy of the retinal pigment epithelium (CHRPE).CHRPE, which
Retinal Lesions and APC MutationsNalanzano et al.
occurs in up to 60% of patient^,^-^ is the most frequent
extracolonic manifestation of the disease and is widely
used a!; an early clinical marker. However, the sensitivity of CHRPE analysis varies in the literature, probably
due to differences in the criteria utilized to define a
positive ophthalmic exarninati~n.’~~
FAP is caused by germline mutations of the adenomatous polyposis coli (APC) gene.’-’* The APC gene
is comprised of at least 15 coding exons and several
noncoding exons, which can be expressed in various
combinations by alternative s p l i ~ i n g . ~ The
~ ’ ~ ~APC
’~
gene product is a cytoplasmic protein of large size
that contains 2843 amino acids in its most common
form.”” APC proteins can associate to form homodimers i3nd interact with other cytoplasmic proteins, including microtubules and p-catenin. Microtubules
bind 1 he carboxy-terminal region, typically deleted in
mutant proteins, whereas p-catenin, which is necessary for the function of the cell adhesion protein Ecadherin,“ interacts with the region of exon 15 downstream of codon 1014.’6”7The vast majority of the APC
mutations thus far reported in FAP patients result in
premature termination of protein synthesis.I8
There are correlations between position of the
APC mutation and phenotypic manifestations of FAP.
Mutations near the 5’ end of the gene are associated
with an attenuated form of the disease, characterized
by a reduced number of colorectal polyps.’ Mutations
downstream of exon 9 appear to cause the classic form
of FA.P associated with CHRPE.19,20
The APC gene was analyzed in 47 FAP patients
and in 14 “at risk” relatives from 26 kindreds. The
analysis was conducted using a combination of
screening techniques. These included single strand
conformation polymorphism (SSCP) analysis of the
coding region from exon 1 through exon 15, heteroduplex analysis coupled with multiple allele specific
polymerase chain reaction (PCR) for the identification
of 3 common mutations in exon 15,” and the in vitro
synthesized protein assay (IVSP) for the detection of
truncating mutations of exon 15.18,21-23
The 61 subjects
who were analyzed at the molecular level are part of
a larger group of 39 FAP patients and 53 relatives from
31 kindreds subjected to ophthalmoscopic studies using novel criteria designed to allow a standardized assessment of CHRPE positivity. The purpose of this article was to describe and evaluate the results of CHRPE
studies in the light of data derived from molecular
analyses, colonoscopy, and standardized criteria for
the evaluation of ocular lesions.
PATIENTS AND METHODS
Patients
Starting in 1988, 39 affected patients (group I) and 53
relatives at 50% genetic risk of FAP (group II), belong-
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ing to 31 kindreds, were evaluated for the presence of
CHRPE by the same ophthalmologist. The individuals
studied included 53 males and 39 females. The mean
age of Group I subjects was 32.2 years 2 13.9 standard
deviation (SD) (range, 8-62 years); the mean age of
Group I1 subjects was 23.3 years 5 10 SD (range, 245 years). All Group I1 subjects aged older than 10
years (47 of 53) were also screened by colonoscopy.
Molecular genetic analysis of the APC gene was performed on 61 subjects from 26 kindreds (Group I: 47
subjects; Group 11: 14 subjects).
Ophthalmoscopic Examination
CHRPE screening
All individuals received a dilated fundus examination,
including direct and indirect ophthalmoscopy and biomicroscopy with a Goldman lens. All cooperating
subjects were submitted to wide angle fundus photography, using a Canon 60” Fundus Camera (Japan).The
entire retina was photographically documented with
extensive overlapping peripheral photographs. Each
CHRPE lesion was evaluated with regard to size, location, and degree of pigmentation. All subjects younger
than 10 years of age were followed for a minimum
of 3 years, to evaluate a possible new appearance or
progression of the lesions.
Diagnostic criteria
CHRPE lesions were classified in 3 morphologic
classes: 1) very small (< 100 pm in greatest dimension), hyperpigmented areas (“spot lesions”) with well
defined margins and no halo, localized in the midperiphery (Fig. la); 2) round, depigmented, or hyperpigmented areas ( < I disk in greatest dimension), generally located in the midperiphery (Fig. Ib); and 3) areas
located at the posterior pole or along the main vascular
arcades, with two different aspects: typical hyperpigmented or hypopigmented, oval areas surrounded by
a depigmented halo (“leaf-like” lesions) (Fig. lc); or
large (>1 disk in greatest dimension), round, variably
pigmented areas with regular or scalloped margins
(Fig. Id).
For scoring purposes, each CHRPE lesion was assigned a value ranging from 1 to 3, according to the
class (class 1: score 1; class 2: score 2: class 3: score
3). The individual CHRPE score was calculated by adding the relative values determined for each eye. Fundoscopy was considered positive when the individual
CHRPE score was 23. Using this cutoff level, lesions
of the type observable in normal subjects were considered of significance only if m ~ l t i p l e . ~
Identification of APC Mutation
One propositus from each FAP kindred was initially
subjected to molecular analysis to identify mutations
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CANCER December 1,1996 / Volume 78 / Number 11
FIGURE 1. Examples of congenital hypertrophy of the retinal pigment epithelium
(CHRPE) lesions. (a) Small (< 100 p m in greatest dimension), hyperpigmented area of
CHRPE with well defined margins, localized in the periphery (“spot” lesion), score = 1.
(b) Oval, irregularly pigmented lesion with greatest dimension < 1 optic disk, localized
just below the main vascular archade, score = 2. (c) Typical, oval pigmented lesion
surrounded by a depigmented halo (“leaf-like” lesion), score = 3. (d) Two large (>1
optic disk in greatest dimension), round, irregularly pigmented lesions with regular margins, score = 3 (for each lesion).
in the APC gene. In the kindreds in which mutations
were detected, mutational analysis was extended to
“at risk” relatives. The screening of APC mutations
was performed by three independent methods. The
first method was designed to identify three frequently
occurring deletions of the APC gene at codons 1061,
1068, and 1309, and was comprised of heteroduplex
analysis on agarose minigel (HAAM), coupled with allele specific multiple PCR. This diagnostic strategy was
previously described in detail.” The second method
Retinal Lesions and APC MutationsNalanzano et al.
2403
FIGURE 1. (continued)
was comprised of SSCP analysis followed by PCR direct
sequencing. The coding sequence of the APC gene,
spanning exons 1 through 15, was amplified by PCR
and subjected to SSCP as des~ribed."-'~The nucleotide sequence of amplified genomic DNA was directly
determined using 35SdATP as the label.'4 The third
method was comprised of IVSP, as described by Powell
et a1." Because RNA samples from FAP patients were
not available, the latter analysis was limited to exon
15, which contains approximately 60% of the coding
sequence of the APC gene. Genetic analyses, either
positive or negative for the presence of APC mutations,
were always repeated using samples deriving from
new DNA extractions. Duplicate experiments confirmed the results.
Statistical Analysis
Ophthalmoscopic data were analysed by the chisquare test. Confidence limits were calculated to estimate the possible range for sensitivity, specificity, and
predictive values. The Mann-Witney U test was used
to analyze correlations between CHRPE scores and positions of the mutations.
CANCER December 1, 1996 / Volume 78 / Number 11
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* Kindreds with known APC mutation
#Kindreds with undetermined APC mutation
FIGURE 2. Distribution of congenital hypertrophy of the retinal pigment epithelium (CHRPE) scores in Group I (A)and Group I1 (0)
of patients.
Subjects of the same kindred are vertically arranged. The number of subjects with the same CHRPE score is shown inside the symbols.
RESULTS
CHRPE Analysis
Examples of the application of the authors’ scoring
method to the evaluation of CHRPE lesions are shown
in Figure 1. Twenty-nine of 39 Group I subjects (74%)
from 29 families and 16 of 53 Group I1 subjects (30%)
from 22 families had a CHRPE score 2 3. Three young
subjects of Group 11, aged 2, 8, and 9 years, respectively, had only 1 or 2 “spot” lesions each at the first
eye examination and were considered negative for
CHRPE. A second fundoscopy was performed at 5, 13,
and 14 years, respectively. The first subject showed 3
typical “leaf-like’’lesions, 1 “spot,” and 1 Class 2 lesion (total score = 12); the second subject showed 1
“leaf-like’’lesion, 6 “spots,” and 1 Class 2 lesion (total
score = 11); the last subject exhibited 5 “spot” lesions
(total score = 5). Figure 2 illustrates the distribution
of the CHRPE scores in both group of patients, within
and among the 31 families examined, and shows the
extreme variability in CHRPE expression (scores between 0 and 16).
Colonoscopy revealed polyps in 20 of 47 Group I1
TABLE 1
CHRPE Status and Results of Colonoscopy in 47 Group I1 Subjects
~~
Polyps
CHRPE positive
(no.of subjects)
CHRPE negative
(no. of subjects)
Present
15
5
Absent
1
26
CHRPE coneenital hwertroohv of the retinal oiament eoithelium
subjects examined (42%). Fifteen of these 20 subjects
were positive for CHRPE, with a mean score of 8.4 ?
4 SD (range, 3-15). Five Group I1 subjects negative for
CHRPE were positive for polyps at colonoscopy (Table
1). Only one of the 27 Group I1 subjects who was negative at colonoscopy was positive for CHRPE. This subject exhibited a depigmented area < 1 disk greatest
dimension in the right eye and a “spot” lesion in the
left eye and thus had a CHRPE score of 3. Ten of 39
FAP Croup I subjects were negative for CHRPE. The
distribution of the CHRPE scores in the 59 subjects
Retinal lesions and APC MutationsNalanzano et at.
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H H H-IH-t-
Subjects with colorectal
polyps (group I + group 11)
Subjects without colorectal
Polyps (group 11)
Distribution of the congenital hypertrophy of the retinal pigment epithelium (CHRPE) scores in subjects positive ( + : Group I; 0 : Group
!I) and negative (U) for colorectal polyps at colonoscopy.
with colorectal polyps (39 in Group I and 20 in Group
11) and in the 27 Group I1 subjects negative for polyps is
shown in Figure 3. The difference in CHRPE positivity
between subjects negative and positive for polyps is
significan.t ( P < 0.001) (Fig. 3 ) .
The cumulative sensitivity of CHRPE was 72.88%
(95% confidence limits [CL],59.73-83.63%; P < 0.05)
and the specificity was 96.29% (95% CL, 81.02-99.90%;
P < 0.05). The positive and negative predictive values
were 97.72% (95% CL, 87.97-99.94%; P < 0.05) and
61.90% (95%CL, 45.63-76.42%; P < 0.051, respectively.
Molecular Analysis of the APC Gene
Germline mutations of the APC gene were identified
in 17 unrelated FAP kindreds using 3 independent
screening methods (Table 2). A total of 34 carriers of
germline APC mutations were identified in the 17 kindreds (Fig. 4). The frequently occurring deletions of
the APC gene at codons 1061, 1068, and 1309 were
identified in 9 pedigrees (GD-3, GD-4, GD-5, GD-6,
GD-7, GD-8, GD-9, GD-10, and GD-11) using HAAM
coupled with allele specific multiplex PCR." In 6 additional kindreds (GD-1, GD-2, GD-12, GD-13, GD-15,
and GD-17), mutations were detected using SSCP coupled with direct s e q u e n ~ i n g . ~Finally,
~ ~ ' ~ , ~in~ 2 kindreds (GD-18 and GD-19), in which SSCP analysis
failed to detect the germline mutations, IVSP revealed
the presence of truncated APC alleles that were further
characterized by sequence analysis (Table 2). Mutations at codons 367,876,995,1112,and 1075 are novel
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CANCER December 1, 1996 / Volume 78 / Number 11
TABLE 2
Summary of Germline Adenomatous Polyposis Coli Gene Mutations
Detected in 17 Familial Adenomatous Polyposis Kindreds
Kindred
Type of mutation
Codon
Exon
GD- 1
GD-2
GD-3, GD-5, GD-6,
GD-7, GD-10,
GD- 1I
GD-4
GD-8, GD-9
GD-12
GD-13
GD-15
GD-17
GD-18
GD-19
1 bp deletion
3 bp deletion
1324a
437“
15
9a (donor site)
5 bp deletion
4 bp deletion
5 bp deletion
1 bp deletion
Tyr-Stop
Arg-Stop
2 bp deletion
Arg-tStop
Cys-tStop
130ga
106V
1061a
1112
1075a
1114
367
876
995
15
15
15
15
15
15
9
15
15
bp: base pair.
aPpreviouslvoublished in references 21-23 and 25.
in patients with mutations in exon 15 than in patients
with mutations in exon 9 (Mann-Witney U test; P <
0.01) (Fig. 5).
No differences in CHRPE scores were apparent in
carriers of mutations in exon 15 occurring upstream
or downstream of codon 1014 (2 and 26 subjects, respectively). The 2 patients with mutations before codon 1014 had a CHRPE score of 6 and 9, respectively;
the mean CHRPE score of the patients with mutations
after codon 1014 was 8.15 5 3.8 SD (range 3-16).
FAP carrier status was ruled out in only 1 of the
16 Group I1 “at risk” subjects with positive CHRPE
scores. This was a 38-year-old individual (GD 1-12)
who had a depigmented area in the right eye and a
spot lesion in the other eye, with a consequent CHRPE
score of 3, which corresponds to the cutoff level established in this study. Subject GD 1-12 tested negative
for the APC mutation responsible for the disease in his
kindred and was negative for polyps at colonoscopy.
DISCUSSION
germline variants of the APC gene. The deletion of two
base pairs at codon 367 is located in the portion of
exon 9 that undergoes alternative splicing and is expected to be present only in a fraction of transcripts
from the allele bearing this deletion (manuscript in
preparation). All mutations are predicted to result in
truncated APC proteins.
CHRPE Score and Position of APC Mutations
Mutations in exon 9 occurred at the site used in both
forms of alternative splicing of the exon (codon 437)
in one kindred (3 patients), and in the portion of exon
9 that undergoes alternative splicing (codon 367) in
the other kindred (3 patients). None of the patients
with the mutation in the alternatively spliced region
had a positive CHRPE score. Only 1 of the 3 patients
with mutations at codon 437 (GD 2-71) was positive
for CHRPE (score = 6).All carriers of mutations in exon
15 were positive for CHRPE; however, the individual
CHRPE scores varied both among and within the 15
kindreds examined, irrespective of the mutation site
(Fig. 4). In particular, the CHRPE score varied from 3
(determined in 3 patients with mutations at codons
1112, 1114, and 1309, respectively) to 16 (determined
in 2 patients, from 2 different families, with mutations
at codon 1309). The same variability (CHRPE score:
range 3-16) was also observed in the group of 12 patients from 6 kindreds with mutations at codon 1309
and in patients belonging to a single kindred (GD-3)
bearing the 5 base pair deletion at codon 1309. Intrafamilial variability was also significant in patients with
mutations at codon 1324 (from kindred GD1) (Fig. 4).
The mean CHRPE score was significantly higher
Not infrequently, otherwise normal individuals may
have one or two areas of CHRPE, usually appearing as
small (<O. 1 disk greatest dimension), unilateral, hyperpigmented lesion^.^^".^ In normal subjects, such lesions are usually < 4 and are rarely bilateral?’ There
is no agreement on the number and the type of CHRPE
lesions meeting clinical significance. Usually, only the
presence of multiple areas of CHRPE, upward from a
minimum varying from 2 to 4, is considered indicative
of FAP.z7-z9In FAP patients, it has been shown that
small, round, pigmented dots are significantly correlated in number with age,6but there is no evidence of
any variation with age for the typical large oval pigmented lesion^.^ In this study, an increase in the number of “spot” areas and the appearance of new, large
and typical lesions was observed in three young subjects who exhibited only one or two small pigmented
“spot” areas of CHRPE at their first ophthalmic examination. The possibility that the first eye examination
was unreliable due to the young age of the subjects
appears unlikely, at least for two subjects whose entire
retina was photographically documented. However,
ophthalmology may not detect all lesions, because
Traboulsi et al. identified 70 areas of CHRPE at autopsy
in the eyes of a FAP patient with only 24 lesions previously described at f ~ n d o s c o p yTherefore,
.~~
the authors could suspect the growth of already present lesions still not detectable by direct ophthalmology.
The presence of multiple areas of CHRPE (regardless of size) in some controls and in many subjects
with hereditary nonpolyposis colorectal carcinomazg
indicates that the number of lesions per se is not always a reliable sign of FAP. Therefore, in several stud-
Retinal Lesions and APC MutationsNalanzano et al.
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FIGURE 4. Correlation of individual congenital hypertrophy of the retinal pigment epithelium (CHRPE) scores with the site of the adenomatous polyposis
coli gene mutation (codon) in 35 familial adenomatous polyposis patients from 17 kindreds (GD-1 through 13, GD-15, GD-17, GD-18, and GD-19). The
order of presentation of individual patients follows the number of the codon site of the mutation.
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ies, morphologic or dimensional criteria were combined with the number of lesions in the attempt to
define clinically reliable methods for the assessment
of positive ophthalmic examinations. According to
Morton et al., at least 2 of the following 3 criteria
should be met: 1) the presence of bilateral lesions; 2)
I
I
the presence 2 3 lesions; and the 3 ) presence of any
lesion > 0.3 optic disk in greatest dimension." Other
authors propose that only oval pigmented areas with a
surrounding depigmented halo should be considered
pathognomonic of FAP and therefore diagnostically
reliable per se.4331The authors attributed a score of 3
2408
CANCER December 1,1996 I Volume 78 / Number 11
at the cutoff level to each lesion of this type and to
each variably pigmented lesion > 1 disk greatest dimension, a score of 2 to each variably pigmented lesion < 1 disk greatest dimension, and a score of 1 to
each small, pigmented “spot” lesion. Therefore, lesions of the type observable in control subjects were
considered significant only if m ~ l t i p l eThe
. ~ only falsepositive subject detected in this study presented with
a depigmented area and a “spot” lesion with a CHRPE
score of 3, at the borderline. Depigmented areas are
not infrequently observed in FAP patients whereas
they are very rare in subjects at low risk of FAP,4so
that it was reasonable to consider this subject positive
for CHRPE. However, it is also reported that both depigmented and “spot” lesions are of limited diagnostic
value when not associated with other types of
CHRPE.4,32Moreover, a misleading phenotype with
borderline CHRPE has been described.33 Otherwise,
the current scoring method allowed a high specificity,
comparable to that reported by Traboulsi et al. who
consider only the presence of multiple lesions (> four)
and/or bilateral lesions to be significant.**The sensitivity was also in agreement with that reported for
other method^^-^ and even with that of authors who
consider the presence of any lesion significant.’ Moreover, the scoring system allows an immediate comprehension of the patient’s CHRPE status (positive, negative, or borderline) and makes the description of number and type of lesions unnecessary, replaced by
numbers. Thus, although scores within the lower
range of positivity may not always indicate a FAP carrier status, the authors believe their scoring system
is an easy and effective method for the uniform and
comparable evaluation of CHRPE status in FAP patients and in their “at risk” relatives.
Recent studies indicate that there are correlations
between position of the APC mutation and the presence of CHRPE. In particular, it has been reported that
CHRPE is associated with mutations located downstream of exon 9.19,20The region of exon 9 corresponding to the boundary between CHRPE positive and
CHRPE negative mutations has not been clearly defined. Caspari et al. reported the presence of CHRPE
in approximately 50% of FAP patients with mutations
at codons 413 and 438 in exon 9.34Furthermore, it has
been recently observed that patients with mutations
beyond codon 1444 have a high prevalence of desmoid
tumors but lack ophthalmic lesions.34This suggests
that APC mutations introducing stop signals between
exon 9 and codon 1444 of exon 15 result in the expression of protein products that interfere with the proliferative control of retinal pigment epithelium. Such interference might be related to a dominant-negative
effect of the truncated proteins, leading to the inacti-
vation of other proteins binding to the APC gene prodUCt,17,35 or to a reduced stability of APC proteins expressed by alleles carrying mutations in the 5’ region
of the gene.3”
The APC mutations detected in the patients analyzed in this study were comprised within the region
of the gene ranging from codon 367 in exon 9 to codon
1324 in exon 15. The finding of mutations in this region
of the gene might in part be related to the technical
approach. The coding region of the APC gene from
exon 1 through exon 15was screened by SSCP analysis.
However, because of the unavailability of RNA Samples, only exon 15, which comprises approximately
60% of the coding region of the APC gene, could be
analyzed by IVSP. In this region, IVSP detected two
mutations that had escaped detection by SSCP. Therefore, it is possible that some patients with undetermined APC mutations characterized for CHRPE lesions
have mutations in the region of the APC protein encoded by exons 1-14. Alternatively, there may have
been large deletions of the coding region of the gene
that escaped detection by SSCP. The current study
confirms that patients with mutations in exon 9 tend
to have attenuated CHRPE expression. In fact, only 1
of the six patients with mutations in exon 9 presented
with a CHRPE score above the cutoff level. Interestingly, this patient had a mutation that was expected
to completely abolish the correct splicing of exon 9.
In contrast, the 3 patients with mutations in the portion of exon 9 that undergoes alternative splicing had
a CHRPE score < 3. All the patients with APC mutations in exon 15 (between codons 876 and 1324) had
CHRPE scores 2 than 3. This notwithstanding, there
was extreme variability in the CHRPE scores associated with mutations in exon 15, irrespective of the
position of the mutation. Even patients from the same
kindred showed significant differences in CHRPE
scores (Fig. 4). As reported in other ~ t u d i e s , ~ , the
* ~ ,5~ ’
base pair deletion at codon 1309, known to be associated with severe colorectal disease, accounted for the
highest number of FAP patients. It is particularly intriguing that the CHRPE scores associated with this
deletion exhibited the highest range of variability documented in the current study, even within the same
kindred. As reported by Wallis et al.,” no differences in
CHRPE expression were found in carriers of mutations
expected to determine the deletion or preservation of
the catenin-binding
In conclusion, the
current study observations suggest that the site of mutation in the APC gene is not the only factor responsible for the variable expression of CHRPE in FAP patients.
Nevertheless, because ophthalmologic examination is a direct, noninvasive, and inexpensive test,
Retinal Lesions and APC MutationsNalanzano et al.
CHRPE maintains its role in FAP screening, especially
in families with known AFT mutation. Moreover, in
newly diagnosed FAP kindreds, CHRPE expression
could direct the mutation analysis on specific regions
of the APC gene.
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Spirio L, Olschwang S, Groden J, Robertson M, Samowitz W,
Joslyn G, et al. Alleles of the APC gene:an attenuated form
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Giardiello FM, Krush AJ, Petersen GM, Booker SV, Kerr M,
Tong LL, et al. Phenotypic variability of familial adenomatous polyposis in 11 unrelated families with identical APC
gene mutation. Gastroenterology 1994;106:1542-7.
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