Download Carcinomas with DNA Mismatch Repair Deficiency

Carcinomas with DNA Mismatch
Repair Deficiency
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Mohammad Hossein Sanei
Department of pathology
Isfahan university of medical science
CRC: HNPCC
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Autosomal dominant.
Mutation in the mismatch repair genes.
High pentrance (around 75%).
Predilection for right side.
Early age of onset.
Higher incidence of synchronous & metachronous tumors.
6% of CRC.
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Inactivation of DNA mismatch repair plays
an important role in the pathogenesis of 10%
to 15% of colorectal cancers.
Although some of these are due to hereditary
alterations in one or more of the mismatch
repair genes (see Hereditary Non polyposis
Colorectal Cancer, later),
the majority (about 90%) are sporadic, due
primarily to promoter methylation, and
silencing, of transcription of MLH1
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characteristic MSI-H genetic alteration,
these cancers reveal a variety of
specific clinical and pathologic associations
of importance to surgical pathologists
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Sporadic mismatch repair–deficient MSI-H
colorectal cancers show a marked female
predominance (about 4 : 1), increase in
prevalence with age, and are particularly
common in women older than 70 years.[114]
Most (about 90%) sporadic MSI-H
colorectal cancers are located in the right
colon, where they constitute about 50% of all
right-sided cancers in women older than 70
years
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Precursor lesions often reveal characteristics
of sessile serrated polyps (sessile serrated
adenoma) in sporadic cases,
and conventional adenomas in hereditary
cases, such as HNPCC.[116]
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MSI-H cancers are also more likely to be
multiple, have a
polypoid or exophytic growth pattern, and show
sharply
circumscribed pushing margins and marked
grossly visible
necrosis.
Furthermore, MSI-H tumors are more likely to
show
mucinous or
signet ring cell features, and
microglandular differentiation
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Of note, the association between these
histologic features and DNA mismatch
repair deficiency is high even when less than
50% of the tumor reveals these features.
Therefore, the sensitivity and specificity of
these histologic features for the diagnosis of
MSI-H cancer vary according to the specific
criteria used
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The presence of a component of any one of
the associated histologic variants
(mucinous,
signet ring cell,
microglandular, or
undifferentiated) or of a
prominent tumor-infiltrating lymphocytic
response provides a relatively sensitive
threshold to identify possible MSI-H
cancers.
Pathologic Features of High-Frequency Microsatellite Instability Colorectal Carcinomas
Pathologic Feature
Location in right colon
Positive Predictive Value
(%)[*]
32
MSI-H (%)
90
82
13
MSI-L/MSS (%)
34
54
5
15
5
35
22
7
36
13
38
14
25
28
13
0.4
3
8
21
Pushing margin
15
Crohn's-like reaction
49
“MSI-H” by pathologist 49
3
8
36
11
55
Exophytic/polypoid
Signet ring cell
component
Mucinous type (>50%
extracellular mucin)
Mucinous component
(>10% extracellular
mucin)
Cribriform pattern
Poor differentiation
Medullary type
Medullary component
(>10%)
Lymphocytosis
21
31
34
71
59
25 (NS)
19 (NS)
44
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The character of host lymphoid response is
another feature that shows a striking
difference between MSI-H and MSS cancers
(Fig. 23-16).
The presence of tumor infiltrating
lymphocytes is highly predictive of MSI-H
cancer, particularly when present in
combination with an undifferentiated tumor
(which may or may not be of classic
medullary subtype
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In one study, the presence of at least
5 tumor-infiltrating lymphocytes per 10 highpower fields had a sensitivity of 93% and a
specificity of 62% for the detection of MSI-H
colorectal cancer.[122]
A number of complex algorithms have been
proposed in an attempt to achieve a high
degree of specificity for the presence of MSIH in any single colon carcinoma.
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Overall, compared with MSI-H HNPCC–
associated colon cancer,
sporadic MSI-H cancers are also more
frequently heterogeneous,
poorly differentiated,
mucinous,
proximally located, and,
as mentioned earlier, associated with a more
prominent tumor-infiltrating lymphocyte
reaction
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Clinically, patients with MSI-H colorectal
cancers are more likely to present at an
advanced T stage, but are less likely to show
regional lymph node metastases.
ESSENTIAL
PATHOGENETIC
ISSUES
 It
is now known that about 50% of
HNPCC cases (defined by
Amsterdam I/II criteria) are
caused by hereditary defects in one
of four DNA mismatch repair
genes[210–212] (Table 23-18).
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Mismatch repair genes function within a
molecular complex;
MSH2, and its binding partners MSH3
and MSH6, mediate recognition of the
DNA mismatch.
 Subsequent recruitment of MLH1, and
its binding partner PMS2, and other
enzymes, leads to excision,
repolymerization, and finally ligation of
the repaired strand of DNA.
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HighFrequency
Microsatellite Gene
Instability
Mutation
Yes (50%)
MLH1 (39%)
MSH2 (38%)
MSH6 (11%)
PMS2 (7%)
Unknown (5%)
No (50%)
Yes (as above; 10%)
Unknown (90%
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Germline DNA mismatch repair gene
mutations are heterozygous, and involve
only one allele.
Small frameshift mutations are the most
common and result in premature protein
truncation, followed by nonsense mutations
and larger genomic deletions.
Some patients with HNPCC harbor point
mutations that result in amino acid
substitutions at functionally critical residues
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Loss of DNA mismatch repair in tumor cells
leads to a dramatic increase in the rate of
mutations, which is characterized by
frameshift mutations in repetitive DNA
sequences known as microsatellites; this is
referred to as microsatellite instability.[113],
[218]
 Rare
patients have also been
described who harbor bi allelic
inherited mutations in a single
[
mismatch repair gene.
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These individuals have functionally
defective DNA mismatch repair in
all somatic tissues, which results in a
severe, clinically distinctive phenotype
(Lynch III) characterized by
very early onset (teens and early twenties)
and a
preponderance of hematologic and
brain tumors, in addition to the usual types
of Lynch syndrome–associated cancers
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DNA mismatch repair is also defective in
about 12% of all colorectal cancers in the
absence of a germ line DNA mismatch
repair gene mutation.
In the majority of these cases, MLH1
expression is silenced in tumor cells by
methylation of CpG islands in the promoter
region of the gene.
PATHOLOGIC
FEATURES
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Compared with colorectal cancers with
MLH1 methylation,
HNPCC-Lynch cancers are more likely to
occur in
men, present at a
younger age (generally less than 60 years),
be associated with prior colorectal cancers
or Lynch II cancers,
and be located in the left colon or rectum
(about one third overall).
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HNPCC-Lynch cancers are also typically
associated with a
residual precursor adenoma, compared with
a
high association with serrated polyps in
cancers that develop in the MLH1
methylation pathway
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When histologic features are considered, one
study also found that lymphocytic
infiltration and tumor budding are more
common in HNPCC-Lynch cancers,
whereas mucin secretion, poor
differentiation, tumor heterogeneity, and
glandular serration were all more common
in MLH1 methylated cancers
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In a more practical approach, one study
attempted to determine optimal pathologic
features that can be used to help identify
likely HNPCC-Lynch cancers before 60
years of age.
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Using factors such as patient age, anatomic
site, and four pathologic features (
histologic type,
grade,
Crohn's-like lymphoid reaction, and
tumor-infiltrating lymphocytes),
an “MsPath” (microsatellite instability by
pathology) score is generated that can be
used, on a sliding scale, to help predict the
probability that a particular cancer is MSI-H
ANCILLARY STUDIES USED
BY PATHOLOGISTS
Microsatellite Instability Testing
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Frameshift mutations in microsatellites can
be identified by extraction of DNA from
both normal and tumor tissue (usually
paraffin-embedded tissue),
amplification of selected microsatellites by
PCR, and analysis of fragment size by gel
electrophoresis or an automated sequencer
(Fig. 23-28).
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Bethesda consensus–defined panel of
microsatellites[222] or a revised panel that
uses more mononucleotide markers.[223]
The sensitivity of the revised panel of
microsatellite instability testing is at least
90%
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Only occasional tumors from patients with
known pathogenic mismatch repair gene
mutations are MSS.[223]
The specificity of the revised panel is also
very high.
The absence of mutations in up to 20% of
HNPCC families with MSI-H tumors is
believed to be due to a failure to detect
unusual germ line variants
TABLE 23-19 -- Bethesda Criteria for
Microsatellite Instability
Loci with Microsatellite
Instability[*]
Classification
≥40%
High-frequency
microsatellite instability
10%-30%
Low-frequency
microsatellite instability
0%
Microsatellite stable
Mismatch Repair mmunohistochemistry
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The DNA mismatch repair proteins are
ubiquitously expressed in normal human
tissues, particularly proliferating tissues, and
nuclear expression in crypt epithelium and
lymphocytes serves as an internal positive
control for stain quality.
In the setting of HNPCC, most hereditary
and second-hit tumor alterations are
inactivating chainterminating mutations.
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As a result of instability of the truncated
mRNA transcript, or the protein product,
these mutations are associated with
complete loss of immunohistochemically
detectable mismatch repair protein in
tumors[224–226] (Fig. 23-29).
Immunohistochemical testing is typically
performed using antibodies to the four
genes that harbor the most common known
HNPCC mutations: MLH1, MSH2, MSH6,
and PMS2.
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During normal DNA mismatch repair
activation, MLH1 recruits its binding
partner PMS2 to the site of DNA repair.
If MLH1 is mutated (as in HNPCC) and
lost from the DNA mismatch repair
complex, then PMS2 will also be absent
from the repair protein complex.
Therefore, mutation and loss of MLH1
protein is also usually accompanied by loss
of PMS2 expression.
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The sensitivity of DNA mismatch repair
immunohistochemistry for identifying loss
of protein expression in patients with
HNPCC with known mutations is at least
90%.
Immunohistochemical staining is often
intact in tumors from patients with
functionally pathogenic point mutations
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The specificity of loss of protein expression
for an underlying mismatch repair defect is
virtually 100%, although up to 10% of these
tumors are, in fact, MSS on microsatellite
instability testing.[225]
TABLE 23-20 -- Interpretation of DNA Mismatch Repair Immunohistochemistry
MLH1
+
−
PMS2
+
−
MSH2
+
+
MSH6
Interpretation
+
Intact DNA
+
MLH1
mismatch repair; or
rare germline point
mutation with
intact
immunohistochemis
try; or other gene
methylation
silencing; or MLH1
germline mutation
(HNPCC)
+
+
+
+
−
+
−
+
+
−
+
−
MSH2 germline
mutation
(HNPCC)
PMS2 germline
mutation (HNPCC)
MSH6 germline
mutation (HNPCC)
BRAF MUTATION TESTING
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Activating point mutations in the oncogenic
signaling protein KRAS are present in about
40% of colorectal adenomas and
carcinomas, and are considered a
prototypical genetic alteration in these
neoplasms.
Recently, mutations in another member of
the same signaling pathway, BRAF, were
identified exclusively in colorectal cancers
with wild-type KRAS genes.[228]
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Interestingly, BRAF mutations are nearly
exclusively present in serrated pathway
neoplasms, and have not yet been reported
in HNPCC-Lynch syndrome tumors with a
known mismatch repair defect.[230]
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Because of pathway specificity, some
centers have begun to incorporate BRAF
mutation testing into their diagnostic
algorithms as a means to differentiate
sporadic MLH1-deficient (methylated)
cancers from hereditary MLH1-deficient
cancers with an underlying germline
mutation.[227]
In MLH1-deficient cancers, BRAF mutation
testing has a sensitivity and specificity of
50% to 70% and 100%, respectively, for
somatic MLH1 methylation.[227]
CpG Island Methylator Phenotype
Testing
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A subset of colorectal cancers (about 25%)
have widespread aberrations in DNA
methylation, including promoter silencing of
genes that are important to tumor
biology.[231]
Referred to as the CpG island methylator
phenotype (CIMP), this includes most
sporadic MSI-H cancers with methylation
silencing of MLH1.[
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CIMP testing is a method to detect
abnormal DNA methylation by using a
panel of markers/loci, and has been used in
some studies to differentiate sporadic from
hereditary MLH1-deficient cancers.
Although there has not yet been an
international consensus on the correct
choice of markers for CIMP testing, several
loci have begun to emerge as the most
sensitive and specific for this type of
application.[230]
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The goal of a genetic workup of HNPCC
families is to identify the underlying germ
line mutation.
Confirmation of the germ line mutation
allows for the most accurate treatment and
follow-up recommendations for the patient,
and allows predictive testing to be
undertaken in interested family members.
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The initial approach by most laboratories is
to analyze the complete coding sequence of
the relevant gene or genes (depending on
immunohistochemistry results), as well as a
portion of the intronic regions important to
exon splicing
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Some laboratories use a variety of rapid
screening approaches to find mutations,
whereas others undertake a complete
sequence analysis.
In certain specific geographic/ethnic
settings, screening may also begin with
testing of founder mutations common to
some populations (e.g., the American
founder mutation in MSH2).[
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If mutations are not detected, testing for
mRNA splicing abnormalities and large
genomic deletions is indicated, but this may
not always be available.
One of the biggest challenges in germline
testing relates to the interpretation of point
mutations.