Hereditary Nonpolyposis Colorectal Cancer Workup

Updated: Jan 20, 2020
  • Author: Juan Carlos Munoz, MD; Chief Editor: BS Anand, MD  more...
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Approach Considerations

2015 ACG guidelines on genetic testing and management of hereditary gastrointestinal cancer syndromes

The American College of Gastroenterology (ACG) released the following recommendations for the management of patients with hereditary gastrointestinal cancer syndromes—and they specifically discuss genetic testing and management of Lynch syndrome, familial adenomatous polyposis (FAP), attenuated familial adenomatous polyposis (AFAP), MUTYH-associated polyposis (MAP), Peutz-Jeghers syndrome, juvenile polyposis syndrome, Cowden syndrome, serrated (hyperplastic) polyposis syndrome, hereditary pancreatic cancer, and hereditary gastric cancer [15] :

  • The initial assessment is the collection of a family history of cancers and premalignant gastrointestinal conditions and should provide enough information to develop a preliminary determination of the risk of a familial predisposition to cancer.

  • Age at diagnosis and lineage (maternal and/or paternal) should be documented for all diagnoses, especially in first- and second-degree relatives.

  • When indicated, genetic testing for a germline mutation should be done on the most informative candidate(s) identified through the family history evaluation and/or tumor analysis to confirm a diagnosis and allow for predictive testing of at-risk relatives.

  • Genetic testing should be conducted in the context of pre- and post-test genetic counseling to ensure the patient's informed decision making.

  • Patients who meet the clinical criteria for a syndrome as well as those with identified pathogenic germline mutations should receive appropriate surveillance measures in order to minimize their overall risk of developing syndrome-specific cancers.


Other Tests

When a family fulfills the Amsterdam or Bethesda criteria (see History, the Guidelines subsection), examination of tumor tissue is indicated (even those removed years before). Tests include immunohistochemistry (IHC) testing, microsatellite instability (MSI) testing (usually used as a prescreening test), and DNA analysis (considered unnecessary, expensive, and time-consuming) (see Genetic Testing).


Tumor Testing


Immunohistochemistry (IHC) testing for hereditary nonpolyposis colorectal cancer (HNPCC) or Lynch syndrome uses monoclonal antibodies to show which mismatch repair (MMR) proteins are present in a tissue sample. Antibodies are chemically tagged to produce colored stains when they bind to their target MMRs. Samples of tumor tissue are tested using IHC to assess for the MLH1, MSH2, MSH6, and PMS2 proteins associated with colorectal cancer. The absence of a protein suggests a mutation in the gene that produces it.

An IHC pattern with absent staining for MLH1 and PMS2 and positive staining for MSH2 and MSH6 indicates a mutation in MLH1 (see Table 3 below). [16, 17, 18, 19, 20]

An IHC pattern with no staining for MSH2 and MSH6 and positive staining for MLH1 and PMS2 indicates a mutation in MSH2 (see Table 3, below).

Of the tumors from carriers of a germline mutation in MSH6, 5% have been detected with an IHC pattern compatible with MSH2 mutation. Thus, if MSH2 mutation screening results are negative, experts recommend DNA analysis of MSH6. Loss of MSH6 expression is the predominant cause of mismatch repair MMR deficiency in early-onset CRC. Endometrial cancer has also been associated with MSH6 mutation. [21]

In the fourth row of Table 3, below, the IHC pattern matching a mutation in MSH6 is shown as absent staining for MSH6 and positive staining for the remaining 3 MMR proteins. This demonstrates the same principle as MSH2: If no mutation is identified, DNA analysis of MSH2 may be considered.

In the fifth row of Table 3, below, the IHC pattern matching a mutation in PMS2 is shown as absent staining for PMS2, with positive staining for the remaining 3 MMR proteins.

Compared with microsatellite instability (MSI) analysis, IHC has the additional advantage of indicating the MMR gene that is most eligible for DNA analysis. IHC is especially indicative for MMR mutations that result in truncation of the protein, such as frame shift, splice site mutations, large genomic rearrangements, and mismatch, although IHC is not always diagnostic for mismatch. In this case, the protein may be functionally abnormal but is still detected with IHC.

Table 3. IHC staining findings. (Open Table in a new window)

MMR Mutations

Protein Staining

























MSI analysis

MSI is the hallmark of the defective DNA MMR gene. First described in 1993, MSI is a phenomenon found in colorectal cancer DNA but not in the adjacent normal colorectal mucosa of individuals with MMR mutations. MSI is characterized by the expansion or contraction of short repeated DNA sequences caused by insertion or deletion of repeated units (DNA regions with repeated patterns of base pairs).

More than 90% of HNPCC tumors (including both adenomas and cancers) and 15% of sporadic colorectal cancers exhibit MSI. This test is used to detect failure of the DNA MMR machinery to repair errors that occur during DNA replication. Such failure leads to increased length in the variation of simple, repetitive sequences distributed throughout the genome. The presence of instability indicates impairment in the DNA replication and repair system, which may be caused by mutations in the mismatch repair (MMR) genes.

A standardized panel of 5 markers is used (D2S123, D5S346, D17S250, BAT25, BAT26 [and possibly BAT40 to increase test sensitivity]). MSI can be subclassified as MSI-high (MSH-H), if 2 or more markers are positive or at least 30% of the markers show instability, or MSI-low (MSH-L), if only a single marker is positive or less than 30% of the markers show instability. Tumors with no positive markers are referred to as microsatellite stable (MSS).

Carcinomas in MSH6 carriers, particularly endometrial carcinomas, have been shown to present with an MSS phenotype; therefore, if an MSS phenotype is found, IHC of MSH6 should be obtained. Nearly 90% of hMLH1 and hMSH2 mutations result in the MSH-H phenotype, whereas nearly 10% of hMLH6 mutations may result in the MSI-L or MSS phenotype.

Although most microsatellite regions are located in noncoding regions of the genome, some are located in the coding regions of genes involved in growth regulation (eg, transforming growth factor [TGF]-beta receptor type II) and apoptosis (eg, BAX). Alterations in these important regulatory genes are believed to be responsible for the accelerated rate of malignant transformation observed in HNPCC. [16, 17, 18, 19, 20]

MSI analysis has a sensitivity of 93% in detecting MMR deficiency in carriers of MMR mutation. However, this test cannot be used to predict which of the MMR genes harbors a mutation.


Genetic Testing

For hereditary nonpolyposis colorectal cancer (HNPCC) or Lynch syndrome, germline testing may be used to identify mismatch repair (MMR) gene mutations. A blood sample is taken to identify mutations by sequence, deletion, duplication analysis, or rearrangement analysis. However, genetic testing for mutations in DNA MMR genes is expensive and time-consuming. Therefore, researchers have proposed techniques to identify ideal candidates (patients with cancer who are most likely to be HNPCC carriers). [22, 23, 24] The Amsterdam criteria are useful but do not identify up to 30% of potential Lynch syndrome carriers.

Researchers have combined microsatellite instability (MSI) profiling and immunohistochemistry (IHC) testing for DNA MMR gene expression. They have identified an additional 32% of Lynch syndrome carriers that MSI profiling alone would have missed. Currently, this combined MSI profiling and IHC testing strategy is the most advanced method of identifying candidates for genetic testing for Lynch syndrome. The next step would be to consider performing a blood test to assess for HNPCC or Lynch syndrome genetic mutation.

Genetic testing is not necessary to establish a diagnosis of HNPCC or Lynch syndrome and does not provide a definitive diagnosis. The decision to go forward with genetic testing is complex. Patients should consult a genetic specialist, such as a genetic counselor, to discuss the benefits and risks before undergoing genetic testing.

In 1997, the National Cancer Institute (NCI) Workshop on Hereditary Nonpolyposis Colorectal Cancer Syndrome suggested 5 markers for the evaluation of MSI. The results are classified as high microsatellite instability (MSI-H) if two or more markers are positive.

The American Gastroenterological Association (AGA) published a literature summary of MSI test results that found that the highest percentage of MSI-H test results were found in families that met the Amsterdam criteria I, followed by patients with colorectal cancer who were diagnosed before age 35 years. Identical results were seen for MMR gene testing.

In general, genetic testing should be offered when clinical suspicion of HNPCC is firmly established—that is, clinical criteria have been met. It is generally accepted that patients who fulfill the Amsterdam criteria or the broader Bethesda criteria are candidates for testing.

The new guidelines recommend MSI or IHC analysis of tumors from affected high-risk patients as the preferred initial diagnostic strategy, followed by germline testing for hMLH1 and hMSH2 mutations for those with MSI-H tumors or tumors with a loss of expression of one of the MMR gene products. hMSH6 germline mutation testing should be considered when the tumor tests MSI-H.

Direct germline testing without MSI or IHC analysis remains an option for high-risk individuals if tissue testing is not feasible (eg, affected proband's tumor is unavailable) or if there is a strong suspicion of HNPCC and MSI/IHC when testing reveals MSI-L, MSS, or normal expression of hMLH1 and hMSH2.

Benefits of genetic testing include the following [24] :

  • Genetic test results may allow for a more accurate assessment of cancer risk. If a mutation is identified, genetic testing and early cancer screening can be offered to the patient and other at-risk family members.

  • Vigorous surveillance and management can then be recommended for patients with identified mutations.

  • Patients with negative test results can avoid these rigorous, expensive, and time-consuming surveillance and management measures and simply follow the American Cancer Society's (ACS) recommendations for the general population.

  • Genetic testing involves no physical risk other than that of a routine blood draw.

BRAF testing

The test for the V600E mutation in a BRAF gene helps to distinguish between HNPCC and sporadic cancer. Sporadic loss of the MLH1 protein expression may result from hypermethylation in the MLH1 gene promoter. Therefore, tumors that show loss of the MLH1 protein but neither BRAF V600E mutations nor hypermethylation are suspected of being associated with HNPCC or Lynch syndrome. [25, 10]

Concerns related to genetic testing include the following:

  • Genetic testing can be emotionally difficult regardless of the results. [26] For example, an otherwise healthy adolescent may become troubled from the knowledge that he or she is a gene carrier with a strong likelihood of developing cancer. This information may compromise interpersonal relationships and educational plans, as well as career goals.

  • Once the diagnosis of HNPCC syndrome has been established, cancer surveillance and management recommendations within the context of genetic counseling should be extended to all available first-degree and second-degree relatives.

  • For people with mutations, the costs associated with cancer screening and prevention may not be covered by their health insurance provider.

  • Employer discrimination or insurance provider discrimination based on genetic test results is a possibility. As a result, in New York State, a person's genetic test results cannot be given to anyone else without the written permission of the person who was tested.

Table 4. Netherlands surveillance protocol for carriers of an MMR-gene mutation. (Open Table in a new window)


MLH1, MSH2, MSH6 (males)

MSHG (females)


Colonoscopy, every 1-2 years, starting at age 20-25 years

Colonoscopy, every 1-2 years, starting at age 30 years



Ultrasonography and CA-125, every 1-2 years, starting at age 30-35 years; consider hysterectomy after age 50 years

Upper Urinary Tract*

Urine cytology analysis, every 1-2 years, starting at age 30-35 years, if it occurs 2 or more times in a family

Urine cytology analysis, every 1-2 years, starting at age 30-35 years, if it occurs 2 or more times in a family


Gastroscopy every 1-2 years, starting at age 30-35 years, if it occurs 2 or more times in a family

Gastroscopy every 1-2 years, starting at age 30-35 years, if it occurs 2 or more times in a family

* Those with HNPCC with MSH2 mutations are at an increased risk not only for upper urinary tract tumors but also for bladder cancer.


Difficulties in HNPCC

If the patient's family does not fulfill the Amsterdam criteria, but two tumors with an microsatellite stable (MSS) phenotype are encountered, the Lynch syndrome surveillance protocol is currently recommended. [27, 28, 29] The age at which surveillance should be initiated and the surveillance intervals are the same as those recommended in the Netherlands surveillance protocol for carriers of mismatch repair gene mutation (see Table 3 in the Tumor Testing section).

If the patient’s family is considered at risk and the specific gene mutation has not yet been identified, a negative genetic test result does not mean that the patient is not at risk for hereditary nonpolyposis colorectal cancer (HNPCC). The patient is still considered at risk if a family history of cancer, tumor testing, or genetic testing indicates HNPCC, even if genetic testing does not identify the gene mutation. Researchers have discovered only seven mutations known to cause HNPCC but believe that many more remain to be discovered (see Table 1 in the Pathophysiology section).

One important problem to resolve is compliance with recommended screening procedures because of the patient’s fear and denial, as well as socioeconomic and educational barriers.

If a family does not fulfill the Bethesda criteria, no specific analysis for Lynch syndrome is indicated. This does not, however, exclude a hereditary factor in the development of colorectal carcinoma in a family. For that reason the referral criteria for genetic counseling are broader than with the Bethesda criteria.

Also, it is not always clear, before a family is referred and before the family history and medical records are analyzed, if a family truly fulfills these criteria. Individuals with a first-degree relative with colorectal carcinoma have an increased relative risk of developing colorectal carcinoma compared with the population risk, but the cumulative risk is not higher than 10%. If a first-degree relative was diagnosed before the age 45 years or if an individual has two first-degree relatives with colorectal carcinoma, the risk is increased four- to six-fold (cumulative risk higher than 10%).

For these indications, a colonoscopic examination every 5 years from the age of 45 to 50 years has been recommended. However, the American Gastroenterological Association, US Multi-Society Task Force on Colorectal Cancer, and the ACS recommend a colonoscopy every 5 years from age 40 years or 10 years before the earliest diagnosis if an individual has two or more first-degree relatives with colon cancer, or a single first-degree relative with colon cancer or adenomatous polyp diagnosed at an age < 60 years.


Colon Screening Tests

The major indications for colon screening tests are as follows:

  • Individuals harboring deleterious germline mismatch repair (MMR) gene mutations

  • Individuals whose affected relatives are unavailable for genetic testing

  • At-risk relatives who refuse genetic testing

  • At-risk asymptomatic individuals for whom genetic testing is inappropriate (eg, family members of probands with high microsatellite instability (MSI-H) tumors and noninformative MMR gene testing)

Published recommendations for colorectal cancer screening in hereditary nonpolyposis colorectal cancer (HNPCC) are based on expert and consensus opinion. The goals of screening and surveillance are the same: to reduce mortality through the detection of presymptomatic, early-stage cancers and to reduce the incidence through the identification and removal of precancerous adenomas.

Early studies by Love and Morrissey, [30] Vasen et al, [31] and Mecklin et al [32] demonstrated that screening of asymptomatic, high-risk individuals detects early-stage cancers and advanced adenomas, thus providing indirect evidence of screening effectiveness. More compelling evidence is derived from a controlled trial by Jarvinen et al that involved two cohorts of at-risk individuals from 22 families with HNPCC. One cohort (n = 133) underwent colonic screening with flexible sigmoidoscopy plus barium enema or colonoscopy every 3 years; the second cohort (n = 119) declined screening and served as the control group. After 15 years of patient observation, 8 screened subjects (6%) developed colorectal cancer compared with 19 unscreened subjects (16%; P = 0.014).


In 1997, colonoscopy was added to the guidelines as a screening option. Colonoscopy is considered the preferred screening test in patients with HNPCC. The evidence to support colonoscopy is derived from data that show a decreased mortality rate in patients with colorectal cancer who have undergone colonoscopic adenoma removal. Additionally, colonoscopy screening is cost-effective compared with other screening strategies.

According to the American Cancer Society (ACS) guidelines, in those with a family history, colonoscopy should be offered every 5 years beginning at age 40, or 10 years before the first diagnosed colorectal cancer in the family, to patients with a family history of colorectal cancer or adenomatous polyps in two or more first-degree relatives younger than 60 years. [22, 33]

For patients with a family history of colorectal cancer or adenomatous polyps in any first-degree relatives older than 60 years, or in at least two second-degree relatives at any age, colonoscopy should be performed every 10 years beginning at age 40. [22]

For individuals with, or at increased risk of Lynch syndrome, colonoscopy should be performed every 1-2 years beginning at age 20-25 years (or 10 years before the first diagnosed colorectal cancer in the family), or annually for those who are confirmed mutation carriers. [22] Genetic testing counseling should be offered to these patients as well as to first-degree relatives of patients identified as carrying Lynch syndrome mutations and to those who meet one of the first three Bethesda criteria (see History for these criteria). [22]

Current cumulative data has supported ACS guidelines regarding colonoscopy screening for patients with MLH1 and MSH2 mutations. However, female patients with MSH6 mutations have a lower risk of colorectal cancer. As a consequence, a new guideline has been proposed for this group of patients: colonoscopy starting at age 30 years as opposed to age 20-25 years. [34, 35, 36] To date, no prospective cohorts of MSH6 mutation carriers have been published to confirm the efficacy of this new screening strategy.

After age 40 years, colonoscopies should be performed every 1-2 years. [37] This cost-effective strategy reduces both the incidence of and mortality from HNPCC-associated colorectal cancer.

Barium contrast study

Barium enema has the advantage of allowing visualization of the entire colon. However, there is evidence to suggest that this technique is inaccurate in the detection of small polyps and early cancers and is suboptimal for colorectal cancer screening or surveillance in patients with HNPCC.

In a prospective study comparing the use of double-contrast barium enema and colonoscopy, the barium enema missed 52% of polyps smaller than 1 cm. If barium enema is the only option for screening or surveillance, it should be coupled with flexible sigmoidoscopy. The use of flexible sigmoidoscopy allows visualization of the rectosigmoid, which the barium enema may not depict well because of overlapping bowel loops. Lesions detected on barium enema warrant colonoscopic evaluation.

Flexible sigmoidoscopy

Flexible sigmoidoscopy is not significantly sensitive in individuals with HNPCC mutations, because an estimated two thirds of HNPCC tumors develop on the right side of the colon (proximal to the splenic flexure). Thus, flexible sigmoidoscopy should always be combined with at least barium enema when patients with HNPCC are screened.

Virtual colonoscopy

In virtual colonoscopy, computed tomography (CT) scanning is used to create a 3-dimensional (3-D) image of the air-extended, prepared colon. This procedure has potential as a colorectal cancer screening test in patients with HNPCC.

Virtual colonoscopy does not carry the risks of traditional colonoscopy, such as sedation or perforation. However, if a polyp is detected, a traditional colonoscopy would have to be performed to remove and analyze the polyp. The sensitivity for small polyps has been questioned, and negative virtual colonoscopy results may not be truly negative for the presence of a polyp. This test is not currently recommended as a screening test for patients who carry an MMR gene mutation.


Surveillance for Extracolonic Malignancies

Immunohistochemistry (IHC) and microsatellite instability (MSI) testing may be useful in carriers of a mismatch repair (MMR) gene mutation (see Table 3, under Tumor Testing), as proposed in the Netherlands surveillance protocol.

Women with Lynch syndrome are recommended to have an annual pelvic examination, ultrasonographic examination, and biopsies of the uterus starting at around age 30 or 35 years to screen for uterine cancer. [38]

In at-risk women, an annual screening test for endometrial cancer is recommended, beginning at age 20-30 years. No consensus has been reached on the optimal method of screening: endometrial aspiration (for cytology or histology) or biopsy and transvaginal ultrasonography (see Prophylactic hysterectomy and salpingo-oophorectomy in the Treatment, Surgical Care section).

An annual screening test for ovarian cancer is also recommended. [38] Transvaginal ultrasonography and the ovarian cancer blood test (CA-125) are occasionally used for screening beginning at age 25-30 years, but these techniques are not sensitive and often reveal only cancerous tumors at a later stage. An alternative is to have surgery to remove at-risk body parts before cancer develops. Thus, in addition to recommending endometrial cancer surveillance, it is prudent to recommend to women in families with HNPCC to seek prompt evaluation of abnormal menstrual bleeding, regardless of their last surveillance exam results.

Based on the results of current data, routine upper endoscopic surveillance is of no benefit in Lynch syndrome. However, selective use of upper endoscopy may be justifiable in high-risk kindred. Although there are authors who do not support routine upper endoscopy evaluation in patients with HNPCC, [39] most authorities, including the International Collaborative Group on Hereditary Nonpolyposis Colorectal Cancer, recommend an upper endoscopic procedure every 1-2 years beginning at age 30 years for at-risk relatives with HNPCC-associated gastric cancer. [31, 38, 2, 40, 41, 42, 43]

For example, in the Asian population (eg, Japanese, Koreans, and Chinese), gastric cancer is the second most common cause of cancer-related death; therefore these individuals are probably at a higher risk than other people.

Most of these studies have been done in Japan and China, where the rates of stomach cancer are much higher. To date, the data have been inconclusive as to whether these screening tools affect the stomach cancer prognosis. Therefore, routine stomach cancer screening for the general population is not recommended. However, stomach cancer screening may be recommended for some people who are at a greater risk of developing the disease. People who are considered at risk include:

  • Elderly people with atrophic gastritis or pernicious anemia

  • Patients with partial gastrectomy

  • Patients with the diagnosis of sporadic adenomas

  • Those with FAP

  • Those with HNPCC

  • Immigrant ethnic populations from countries with high rates of gastric carcinoma

  • Even in these groups of people, the impact of screening on death from stomach cancer is not known.

A similar argument can be made for selective surveillance of Lynch syndrome kindred who exhibit small bowel adenocarcinomas. Despite the lack of supporting data, endoscopic surveillance with enteroscopy or capsule endoscopy every 1-2 years may be justifiable in high-risk individuals (Asian population).

For families with a history of urinary tract tumors, upper urinary tract screening tests may be beneficial. These families should undergo urinary tract ultrasonography, cystoscopy, and urinary cytology every 1-2 years beginning at age 30-35 years.

At this time, no specific screening recommendations for hepatobiliary tract cancers exist, and, in general, liver and biliary tract screening tests are not recommended. However, hepatocellular carcinoma is known to be associated with polyposis coli, and children with maternal ancestors who were affected with the syndrome have developed hepatoblastoma. For families with a history of this type of cancer, transabdominal ultrasonography of the biliary tree and liver function tests have been suggested. [44, 45, 46]


Histologic Findings

Adenomas are often villous with components of high-grade dysplasia and exhibit an accelerated rate of malignant transformation. Colorectal cancers in hereditary nonpolyposis colorectal cancer (HNPCC) have a more aggressive histology (increased frequency of poorly differentiated, mucinous, and signet cells).