Case 3 -
Poorly Differentiated and Microsatellite Unstable Adenocarcinoma Arising in a Patient with Lynch Syndrome
Wendy L. Frankel
The Ohio State University School of Medicine
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58 year old man with no personal history of malignancy or other significant medical
history was noted to have two tumors at colonoscopy. The tumors were located in his transverse colon and
Case 3 - Slide 1
Case 3 - Figure 1
Low power view of the tumor with a pushing border and adjacent lymphoid aggregates.
Case 3 - Figure 2
In some areas, the cells form glands and nests.
Case 3 - Figure 3
In other areas, the tumor cells show a medullary pattern. The malignant cells are arranged in a somewhat trabecular pattern and contain vesicular chromatin with nucleoli. There is a lymphoid infiltrate surrounding tumor cells.
Case 3 - Figure 4
High power view of an area with undifferentiated sheets of tumor cells.
Case 3 - Figure 5
The tumor is immunoreactive with MLH1 and PMS2.
Case 3 - Figure 6
MSH2 and MSH6 mark lymphocytes and benign epithelial cells but staining is absent in tumor cells.
Case 3 - Figure 7
The algorithm used at the Ohio State University on all colorectal carcinoma (CRC) patients to identify Lynch Syndrome and microsatellite instability (from Bellizzi and Frankel, Adv Anat Pathol, 2009).
Colorectal carcinoma is one of the leading causes of cancer in the US. Most tumors
(85%) arise through the chromosomal instability pathway but 15% are due to microsatellite instability.
Patients with these tumors have a better prognosis than those with stage-matched microsatellite stable
tumors. Additionally, they do not respond as well to some chemotherapeutic agents (5- fluorouracil).
Many of these tumors are due to hypermethylation of the MLH1 promoter and are sporadic rather than
germline. These tumors typically occur in older (eighth decade) people and may be associated with a
serrated polyp (serrated pathway). There is a smaller subset of microsatellite unstable tumors that are
due to a mutation in one of the mismatch repair genes (MLH1, MSH2, PMS2, MSH6). These tumors occur in
those with Lynch syndrome and are more common in younger patients (fourth or fifth decade). These
individuals have an increased risk of synchronous colorectal carcinomas and other associated malignancies
such as endometrial, stomach, renal pelvis and ureter cancer. Family members also need screening.
Pathological/Microscopic Findings and any Immunohistochemical or Other Studies: The colon contains
polypoid tumors in the transverse colon and rectosigmoid. The rectosigmoid tumor is shown in sections.
At low power, a pushing border is identified. Lymphoid aggregates are present at the edge of the tumor.
The tumor is poorly differentiated and the morphology varies in different areas of the slide. In some
areas, the cells are arranged as glands and nests. Other areas show a medullary pattern with tumor in a
sheet-like to trabecular pattern containing cells with vesicular chromatin, nucleoli and many lymphocytes
surrounding tumor cells. Undifferentiated areas exist. MLH1 and PMS2 immunohistochemical stains marked
the tumor cells while MSH2 and MSH6 stained lymphocytes and benign epithelial cells but not tumor cells.
Microsatellite unstable adenocarcinoma- Lynch Syndrome Microsatellite
unstable adenocarcinoma- sporadic Microsatellite stable adenocarcinoma
Poorly differentiated and microsatellite unstable adenocarcinoma arising in a patient with Lynch Syndrome
Other important history in our patient includes a very significant history of
carcinoma in other members of his family including endometrial, ureter, lung, colorectal, thyroid,
sebaceous adenoma and brain (glioblastoma multiforme). The pathologic findings of synchronous and poorly
differentiated adenocarcinoma with a pushing border, Crohn-like reaction, tumor infiltrating lymphocytes,
and medullary features suggest microsatellite instability. The patient's age and family history are
indicative of Lynch syndrome. The immunohistochemical findings of intact (present) staining for MLH1 and
PMS2 and absent staining for MSH2 and MSH6 suggest an MSH2 mutation which was confirmed with mutational
testing for MSH2 (patient consent necessary). The patient was found to have the most common mutation in
MSH2, the Desai mutation. His family was screened for this mutation; it was found in 4 of 9 siblings, 1
of 2 daughters and 3 others (nieces, nephews). His family is considered to have the Muir-Torre variant
due to the history of sebaceous tumors and Turcot's due to the brother with glioblastoma multiforme. The
family continues to be followed closely.
Review of the Literature/Treatment Options:
Approximately 15% of colorectal carcinomas show
microsatellite instability rather than chromosomal instability. The majority of these tumors are
sporadic and are due to epigenetic silencing of the MLH1 promoter by
hypermethylation. Both sporadic and hereditary microsatellite unstable carcinomas have prognostic and
predictive implications. Microsatellite unstable colorectal carcinomas have a better prognosis than
matched stage microsatellite stable carcinomas, and tumors do not respond as well to 5-fluorouracil based
chemotherapy. Some institutions are using this information to help guide treatment, particularly in
lower stage tumors where the use of chemotherapy is more controversial.
Lynch Syndrome is the most common hereditary form of colorectal cancer. It accounts for 1 to 5% of
colorectal cancers and is due to a mutation in one of the mismatch repair genes manifesting as
microsatellite instability. The syndrome is characterized by early onset (fourth or fifth decade) and
frequently right sided colorectal cancer. However, tumors can occur anywhere in the colon or rectum and
the age range is wide including older patients. Additionally, patients are at an increased risk for
synchronous or metachronous colorectal cancers and other tumors such as endometrial, ureter, and
gastric. Lynch Syndrome follows a dominant pattern of inheritance with a high penetrance. Patients and
families need screening and counseling to help prevent additional carcinomas or identify them at an early
Lynch Syndrome is identified using history, tumor morphology and/or laboratory tests that identify
mismatch repair deficiency in the tumor. Amsterdam II criteria are clinical criteria to identify
families with HNPCC (hereditary nonpolyposis colorectal carcinoma) while the revised Bethesda guidelines
were developed to identify those who should get microsatellite instability testing. In unselected
patients both show limited sensitivities and specificities. Smaller family size and the increased use of
screening colonoscopies and polypectomies also decrease sensitivity of the criteria. Of course, these
clinical criteria are not useful to detect the larger group of microsatellite unstable colorectal
carcinomas due to an epigenetic event. Therefore, additional testing is necessary to identify these
Histologic features can be useful to detect microsatellite unstable colorectal carcinomas. Some of
the most common include mucinous, signet-ring cell, medullary and poorly differentiated features. Tumors
also show a pushing border, a Crohn-like reaction and tumor infiltrating lymphocytes. It is important
to remember that there is significant histologic phenotypic overlap between sporadic and Lynch-associated
microsatellite unstable tumors. Jeremy Jass has argued that the constellation of histologic features
classically associated with microsatellite unstable colorectal carcinomas largely represents sporadic
rather than Lynch-associated tumors. The most useful feature to help differentiate these tumors, when
present, is a serrated background suggesting methylation and the serrated pathway. Table 1 compares the
clinicopathologic features in Lynch Syndrome to sporadic microsatellite unstable colorectal carcinomas.
Table 1: Clinicopathologic Features of Lynch Syndrome-Associated and Sporadic Microsatellite
Unstable (MSI) Colorectal Cancers
|Age ||< 50 yrs
||> 70 yrs
|Sex ||F = M
||F > > M
|Other Lynch associated tumors ||Yes
|Proximal tumor a ||Yes
|Mucinous a ||Yes
|Signet-ring cell ||Yes
|Intraepithelial lymphocytes ||Yes
|Crohn's-like reaction ||Yes
|Pushing border ||Yes
|Poor differentiation a ||Yes
|Tumor budding ||Yes
|Tumor heterogeneity a ||Yes
|Serration/serrated precursor ||No
|Absent MLH1 and PMS2 ||Yes, some
|Absent MSH2 and MSH6 ||Yes, some
|Absent MSH6 or PMS2 ||Yes, few
|BRAF mutation ||No
|MLH1 promoter methylation ||Rare
|Germline mutation in mismatch repair gene ||Yes
a Each of these features was shown to occur significantly less frequently in
"HNPCC-associated" vs. sporadic MSI CRC (Young et al, Am J Pathol, 2001).
Table modified from Bellizzi and Frankel, Adv Anat Pathol, 2009.
In unselected patients, mismatch repair deficiency can be identified by microsatellite instability
(MSI) testing on tumor DNA or by using immunohistochemistry (IHC) to detect absent mismatch repair
proteins. MSI testing is done in a molecular lab and the mismatch repair deficiency manifests as
microsatellite instability. IHC is typically done by performing a panel of MLH1, MSH2, MSH6 and PMS2.
Some laboratories have demonstrated efficacy using a smaller panel. The staining pattern of these
proteins can vary and may be diffuse or focal but must be nuclear to be considered present. Fixation may
affect staining. Absence should be virtually complete in order to consider the protein lost. The
mismatch repair proteins function as heterodimers. MSH2-MSH6 and MLH1-PMS2 pairs act in concert. If the
obligate partner in the pair (MSH2 or MLH1) is missing, neither protein in the pair will be expressed.
Therefore, a mutation or epigenetic silencing of MLH1 will lead to loss of MLH1 and PMS2 protein, while
a PMS2 mutation will only manifest as negative PMS2 staining with intact MLH1. The pattern of loss can
help point towards the gene that is not being expressed by mutation or an epigenetic event. When MLH1
and PMS2 are absent (most common abnormal pattern), this is either due to Lynch Syndrome or sporadic
methylation of the MLH1 promoter and additional testing is necessary. When
MSH2 and MSH6 or MSH6 or PMS2 protein(s) is lost, additional testing is necessary to evaluate for the
high likelihood of Lynch Syndrome.
Both IHC and MSI testing have limitations and choosing the best test may be dependent upon resources
and ease, cost and speed of results at different institutions. Most studies including ours have shown
the MSI and IHC are comparable to identify Lynch Syndrome. MSI requires tumor and normal DNA and
requires sufficient tumor percentage. IHC only requires tumor (since lymphocytes and endothelial cells
can serve as positive controls), and a small number of tumor cells is usually sufficient, but poor
fixation can affect results. IHC does help suggest the likely gene of interest, and therefore, may help
decrease cost. Some argue this comes close to "crossing the line" of germline testing without consent.
The literature has argued convincingly that IHC is not germline testing since it identifies proteins that
can be altered by epigenetic events. Table 2 compares IHC for the mismatch repair proteins to MSI
testing to screen colorectal carcinomas for a mismatch repair deficiency.
Table 2: Comparison of Immunohistochemistry for the Mismatch Repair Proteins versus Microsatellite
||Microsatellite Instability (MSI)
|Cost ||Variable ||Variable ||Neither|
|Training to perform ||Necessary ||Necessary ||Neither|
|Analyte ||Protein ||DNA ||Neither|
|Laboratory ||Most pathology labs ||Need molecular lab ||IHC|
|# tumor cells needed ||Very few required ||Need certain % depending on marker ||IHC|
|Sample ||Tumor only ||Tumor and normal ||IHC|
|"Contamination" by normal may interfere ||No ||Yes ||IHC|
|Suggests mismatch repair gene involved ||Yes ||No ||IHC|
|Turnaround time ||Next day ||2 to 7 days ||IHC|
|"Genetic" debate a ||Yes ||No ||MSI|
|Distinguishes MSI-H, MSI-L, MSS b ||No (MSI-L and MSS similar staining) ||Yes ||MSI|
|Variable results due to tissue fixation ||Yes ||No ||MSI|
a the "genetic debate" questions whether IHC is a genetic test since results suggest the
likely involved gene. There is no debate with MSI testing. We and most publications do not consider IHC
a genetic test.
b Differentiating MSI-H from MSI-L and MSS does not matter for identification of Lynch
syndrome but may matter for prognosis or predictive reasons.
Table modified from Bellizzi and Frankel, Adv Anat Pathol, 2009.
There are multiple algorithms for testing colorectal carcinomas depending on if IHC or MSI testing is
used as the initial screening test. We advocate using Braf mutational
testing (some choose to use MLH1 promoter methylation testing) in those with
absence of MLH1 and PMS2 proteins before embarking on MLH1 germline mutation
analysis. Since this pathogenic mutation has not been described in those with Lynch syndrome, there is
no need to do additional testing for Lynch syndrome on those with this mutation. The majority of
mutations are accounted for by a point mutation resulting in the substitution of glutamic acid for valine
at codon 600 (V600E). The algorithm used at the Ohio State University is show in Figure 7.
It is important to identify microsatellite instability and Lynch syndrome for both
prognostic and predictive information as well as to screen affected individuals and family members in
Lynch Syndrome. Currently this information is not required in the AJCC TNM 7th edition. The CAP
colorectal checklist does include histologic features such as differentiation and histologic type
(mucinous, medullary or signet ring cell among others). Other features suggestive of MSI including
peritumoral lymphocytic response (Crohn-like reaction), intratumoral lymphocytic response
(tumor-infiltrating lymphocytes), and tumor subtyping (including percent mucinous component) are listed
as optional. Other optional items include ancillary testing such as MSI, immunohistochemical stains for
the mismatch repair proteins and Braf testing. Many labs are doing MSI, Braf and immunohistochemical
testing upon request. It is becoming more common for reflexive testing for MSI or immunohistochemistry
in all colorectal carcinomas or at least in those with suggestive patient age and morphologic features.
The best algorithm for testing is laboratory dependent. Communication with the surgeons and
gastroenterologists is vital in whatever algorithm is chosen.
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