Colorectal Cancer

Indications for Testing

Rectal bleeding, melena, hematochezia
Change in bowels (eg, constipation, narrow stools, excessive flatulence)
Abdominal pain or discomfort; may be accompanied by weight loss, fatigue, and/or anemia
Family history of colorectal cancer (CRC)
Clinical symptoms consistent with genetic/hereditary CRC (eg, MUTYH-associated polyposis [MAP], familial adenomatous polyposis [FAP], Turcot syndrome, Gardner syndrome)

Laboratory Testing

Molecular testing

Tissue from metastatic or recurrent colorectal tumor – preferred if present; otherwise, primary tissue is acceptable
Formalin-fixed, paraffin-embedded tissue – preferred for biomarker testing; other specimen testing is not validated

Molecular biomarkers and recommendations for testing (ASCP, CAP, AMP, ASCO, 2017)

Molecular Biomarkers and Recommendations for Testing Based on Guideline from ASCP, CAP, AMP, and ASCO (2017)
Marker(s)	Appropriate Population to Test	Recommended Use
RAS (“expanded” or “extended” RAS) KRAS NRAS Codons 12, 13 of exon 2 Codons 59, 61 of exon 3 Codons 117, 146 of exon 4	Colorectal cancer (CRC) patients being considered for anti-EGFR therapy	Determine eligibility for anti-EGFR monoclonal antibody therapy in patients with metastatic CRC – mutations in KRAS or NRAS are associated lack of response to anti-EGFR therapy
BRAF p.V600	Select CRC patients, including those with metastatic disease	Prognostic stratification Patients with BRAF-mutated CRC have a worse outcome relative to patients with nonmutated tumors Insufficient evidence to recommend BRAF mutational status as a predictive molecular biomarker for response to anti-EGFR therapy
BRAF pV600E	CRC patients with deficient MMR tumors with loss of MLH1	Evaluate for Lynch syndrome risk Presence of BRAF mutation – strongly favors sporadic pathogenesis Absence of BRAF mutation -- does not exclude Lynch syndrome
Mismatch repair status (MMR)	All CRC patients	Prognostic stratification Evaluate for Lynch syndrome risk Presence of MMR protein deficiency warrants further workup Identify patients at high risk for Lynch syndrome
PIK3CA	n/a	Insufficient evidence to recommend testing for therapy selection outside of clinical trials
PTEN (IHC or FISH)	n/a
Sepulveda, 2017

Genetic testing for other CRC syndromes should be based on clinical presentation and family history and in conjunction with genetic consultation
- Refer to NCCN guidelines – Genetic/Familial High Risk Assessment: Colorectal, 2017

Histology

All polyps removed should have histologic examination
- Tissue – gold standard for tumor classification
- Immunohistochemistry (IHC) – identifies mismatch repair (MMR) protein deficiency and can guide selection of genes for confirmatory genetic testing
  - All tumors should be tested by IHC for MMR deficiency (or polymerase chain reaction [PCR] for microsatellite instability [MSI])
  - 15% of sporadic CRCs will have MSI

Prognosis

Prognosis for CRC is dependent upon clinical and tumor factors
- Patient age
- Cancer stage
- Histology
- Depth of penetration of primary tumor
- Number of lymph nodes evaluated and number positive
- Lymphocytic invasion
- Perineural invasion
- Anatomic location – patients with left-side tumors have better prognosis than those with right-side tumors
Markers for prognosis include
- MSI/MMR-deficient tumors – generally more favorable prognosis
- Chromosomal alterations in 8p, 17p, 18p – associated with poor prognosis
- Serum markers
  - Carcinoembryonic antigen (CEA) – measure prior to surgery (NCCN, 2017)
  - Carbohydrate antigen (CA) 19-9 – potential but not validated serum marker
    - Elevation at baseline may predict worse prognosis
  - Circulating tumor cells (CTCs) – independent predictor of progression-free and overall survival

Differential Diagnosis

Diverticulitis
Inflammatory bowel disease
- Crohn disease
- Ulcerative colitis
Irritable bowel syndrome
Hemorrhoids/fissures

Colorectal screening for persons with average risk (beginning at 50 years)

Colorectal Screening for Persons with Average Risk^a (beginning at 50 years)
Organization	Recommendation
USPSTF, 2016	50-75 yrs – Colorectal cancer (CRC) screening recommended; no specific screening test is recommended (screening tests vary according to risks and benefits) 76-85 yrs – consider screening for certain individuals who are healthy enough for colonoscopy, and lack comorbidity that would limit life expectancy Adults in this age group who have never been screened for CRC are more likely to benefit >85 yrs – CRC screening not recommended
AAFP, 2010	50-75 yrs – one of the following screening regimens^b Annual screen for CRC using fecal occult blood test (FOBT) Sigmoidoscopy every 5 yrs combined with high-sensitivity FOBT every 3 yrs Colonoscopy every 10 yrs 76-85 yrs – recommend against routine screening for CRC; however, there may be considerations that support CRC screening in an individual patient >85 yrs – CRC screening not recommended
ACS, 2013	One of the following regimens^b Flexible sigmoidoscopy every 5 yrs Colonoscopy every 10 yrs Double-contrast barium enema every 5 yrs CT colonography every 5 yrs No upper age limit for screening
^aAverage risk – age ≥50 yrs; no history of adenoma or inflammatory bowel disease (IBD); negative family history (not having 1 first-degree or 2 second-degree relatives with CRC, or a clustering of Lynch syndrome-related cancers in the family) ^bPatients with a family member diagnosed with colon cancer <60 yrs should begin colonoscopy screenings every 5-10 yrs starting at age 40 or 10 yrs before the age of the youngest family member diagnosed AAFP = American Academy of Family Practice; ACS = American Cancer Society; USPSTF = U.S. Preventive Services Task Force

High risk patients – begin screening earlier than average risk patients; may need to be individualized based on risk factor
- Risk factors – personal or family history of colorectal cancer (CRC), adenomatous polyp, inflammatory bowel disease, or other inheritable CRC syndrome

Screening strategies

Screening Strategies
Stool-based Tests^a
Tests	Frequency	Description
Fecal immunochemical test (FIT)	Yearly	More accurate than guaiac fecal occult blood test (gFOBT) Can be performed with single specimen
FOBT (eg, gFOBT)	Yearly	50% of confirmed colorectal cancers (CRCs) have a negative FOBT If patient at risk – consider sigmoidoscopy or colonoscopy (even in presence of negative FOBT) Positive FOBT – mandates further evaluation (eg, colonoscopy)
FIT-DNA	Every 1-3 yrs	Multitargeted stool-based DNA testing Reasonable alternative to other stool-based and visualization-based tests (USPSTF, 2016) Emerging screening strategy that combines FIT with testing for altered DNA biomarkers in cells shed into the stool Testing has increased single-test sensitivity for detecting CRC compared with FIT alone
Direct Visualization
Test		Frequency
Colonoscopy		Every 10 yrs
CT colonography		Every 5 yrs
Flexible sigmoidoscopy		Every 5 yrs
Combination Strategy
Test		Frequency
Flexible sigmoidoscopy with FIT		Sigmoidoscopy – every 10 yrs FIT – yearly
Serology Testing
Test		Description
Septin 9 DNA		Biomarker for presence of CRC – high negative predictive values Indicated for individuals ≥50 yrs who have an average risk and no family history of CRC or personal history of polyp removal or CRC Shown to detect cancers in cecum, ascending colon, transverse colon, splenic flexure, descending colon, sigmoid, rectosigmoid junction, and rectum
^aPositive tests require follow-up with direct visualization

Serum carcinoembryonic antigen (CEA)
- Elevated postoperative titer predicts tumor recurrence
- Monitor for changes in concentration from preoperative baseline
  - Stage II or III tumors – measure every 3 months post operation, continuing for 3 years
- Patient with metastatic disease – monitoring may help evaluate treatment response
CT scan
Colonoscopy
Rectosigmoidoscopy (rectal cancer)
Emerging or investigational markers
- Serum circulating tumor cell count (CTC) – in metastatic tumors, used to monitor disease progression and response to therapy
- Serum CA 19-9 – nonspecific marker; may provide additional information in conjunction with CEA
- Deletion 18q – not enough data to recommend use
- 9q22.2-31.2 – promising new marker for hereditary colorectal cancers

UGT1A1 genotyping
- Uridine diphosphate glucuronyl transferase (UGT1A1) is responsible for clearance of irinotecan, a camptothecin analogue used in treatment of advanced colon cancer
- Decreased gene expression may lead to drug toxicity (development of severe neutropenia)
  - 2 gene variants responsible for 98-99% of genotypes in the Caucasian population – *1 and *28 (repeat TA sequence)
- Routine reduction of dose in *28 homozygous is not recommended (Evaluation of Genomic Applications in Practice Working Group, 2014), but may identify patients at risk for adverse events and in need of closer monitoring
  - Selective genotyping based on patient preferences and predicted dosing
    - Higher doses associated with increased risk for certain genotypes that may not occur with lower dosing
    - Patients homozygous for *1 may tolerate aggressive treatment better than patients with the *28 variant
    - Patients homozygous for *28 may require a dose reduction to minimize dose-related adverse events
5-fluorouracil (5-FU) sensitivity – genotyping of DYPD and TYMS
- 5-FU is a fluoropyrimidine drug used in the treatment of colorectal cancer and other solid tumors
- Pharmacogenetic variations in genes such as DPYD and TYMS may contribute to risk of toxicity or altered therapeutic benefits
  - DPYD variant or TYMS variant detected – most variants or mutations are associated with increased risk for 5-FU toxicity
    - Alternative chemotherapeutic agents, therapeutic drug monitoring, altered 5-FU doses, or increased surveillance for adverse drug reactions may be indicated

Colorectal cancer (CRC) is the third most common form of cancer in the U.S. It can be roughly divided into sporadic and hereditary types.

Epidemiology

Incidence – 40/100,000 (2016 U.S. SEER data)
- Sporadic – most common form (~80%)
- Hereditary CRCs
  - Hereditary nonpolyposis colorectal cancer (HNPCC) or Lynch syndrome – accounts for 2-4% of CRC cases in the U.S. (NCCN, 2017)
  - Familial adenomatous polyposis (FAP) – occurs in 1/10,000 live births
    - ~0.5% of total CRC cases
  - MUTYH (formerly known as MYH)-associated polyposis (MAP) – ~1% of Caucasians are predicted to carry an MUTYH gene variant
  - Other inherited syndromes
    - Peutz-Jeghers syndrome (PJS) – 1/200,000
    - Juvenile polyposis syndrome (JPS) – 1/100,000
    - Hereditary diffuse gastric cancer – rare
    - Serrated polyposis syndrome – uncommon
    - Cowden syndrome (PTEN hamartoma syndrome)
    - Li-Fraumeni syndrome – rare
Age
- Sporadic – median is 70 years
- Hereditary – usually <60 years
Sex – M>F

Genetics (Familial Syndromes)

Hereditary syndromes associated with CRC
- Account for >10-15% of all CRCs (NCCN, 2016; ACMG, 2015)
Hereditary CRC types (NCCN, 2017)
- Lynch syndrome (also known as hereditary nonpolyposis colorectal cancer [HNPCC])
  Associated with microsatellite instability (MSI) and mismatch repair (MMR) gene mutations/modifications – usually MLH1, MSH2, MSH6, PMS2
  
  Other genes (MLH3, PMS1, EPCAM, and EXO1) have been identified
- Familial adenomatous polyposis (FAP)
  APC gene variants
  Autosomal dominant inheritance
  
  25% of cases are de novo
  
  Syndromes
  Classic FAP and attenuated FAP forms
  
  Gardner syndrome
  
  Turcot syndrome
- MUTYH (formerly known as MYH)-associated polyposis (MAP)
  MUTYH gene variants
  Autosomal recessive inheritance
  
  2 MUTYH variants, Y165C and G382D, account for 85% of MAP in Caucasians
  
  Biallelic variants appear to create risk for cancer
- Peutz-Jeghers syndrome (PJS)
  STK11 gene variants
  
  Autosomal dominant inheritance
  
  50% of patients do not have this variant
- Juvenile polyposis syndrome (JPS)
  SMAD4 or BMPR1A gene variants
  
  Autosomal dominant inheritance
- Li-Fraumeni syndrome
  TP53 variant
- Cowden syndrome (PTEN hamartoma tumor syndrome)
  PTEN gene variants
  
  Autosomal dominant with high penetrance

Risk Factors

Inflammatory bowel disease (eg, Crohn disease, ulcerative colitis)
First-degree relative with colorectal adenoma or invasive CRC
Tobacco use
Diet high in animal fats/processed meats (Western diet)
Sedentary lifestyle
Patients with metabolic syndrome
Previous adenoma/sessile serrated polyp (SSP)
Previous radiation
Ureterosigmoidostomy – carcinoma can develop ≥15 years post procedure

Pathophysiology

Most CRCs arise from adenomatous polyps
- Villous adenomas transform into adenocarcinomas more frequently than tubular adenomas
- Subset of adenocarcinomas develop from hyperplastic-appearing polyps, especially large, right-sided polyps
- Adenocarcinoma arising in a polyp is considered malignant when it penetrates into the submucosa
Other less-common tumors can occur (neuroendocrine and mesenchymal tumors, lymphomas)

Clinical Presentation

Common signs and symptoms
- Rectal bleeding
- Hematochezia
- Abdominal pain and bloating
- Tenesmus
- Change in stool (narrowing of caliber)
Symptoms vary with tumor location
- Most tumors are located in sigmoid colon and rectum
- Cecum and ascending colon – tumors may be very large before causing symptoms (obstruction)
  - Anemia – a common presenting symptom
- Descending and transverse colon – tumors tend to obstruct and cause annular lesions (apple core or napkin ring) with abdominal pain and bloating
- Rectosigmoid – hematochezia, tenesmus, and narrowing of stool caliber
Sporadic tumors
- Usually single tumors
Clinical presentation and characteristics of inherited syndromes (NCCN, 2017)
- Lynch syndrome
  May have multiple tumors
  
  Median age of onset – 40-60 years (10 years younger than sporadic colorectal carcinoma)
  
  For more information, refer to Lynch syndrome and Lynch syndrome (HNPCC) testing algorithm
- Familial adenomatous polyposis (FAP)
  Classic
  Affected persons develop hundreds to thousands of colon polyps and subsequent colorectal cancer (CRC)
  Often present with multiple tumors
  
  Presence of ≥100 polyps is sufficient for clinical diagnosis (or <100 polyps at younger ages in a family with identified FAP)
  
  Median age of onset – 39 years
  
  Increased risk for other malignancy – thyroid (papillary cribriform-morular variant), medulloblastoma, hepatoblastoma, pancreas, gastric/duodenal
  
  Possible associated findings
  Osteomas – Gardner syndrome
  
  Gastric and duodenal polyps
  
  Dental anomalies
  
  Congenital hypertrophy of the retinal pigment epithelium (CHRPE)
  
  Central nervous system (CNS) tumors – Turcot syndrome
  
  Attenuated
  10-100 polyps (average is 30) – frequently right-sided distribution
  
  Cancer appears at older age than classic FAP – mean age >50 years
  
  Risk of other malignancy similar to classic FAP
  
  Extraintestinal manifestations of classic FAP are rare
- MUTYH (formerly known as MYH)-associated polyposis (MAP)
  100-1,000 polyps (usually <100 polyps)
  
  Median age of onset – 45-59 years
  
  Often indistinguishable from attenuated FAP – may have similar extraintestinal manifestations or serrated polyp syndrome (may have serrated polyps)
  
  Increased risk for malignancy – duodenal cancer
- Peutz-Jeghers syndrome (PJS)
  Hamartomatous gastrointestinal polyps – tend to be fewer than FAP
  
  Perioral melanin pigmentation, which tends to fade when adulthood reached
  
  Increased risk for other malignancy (breast, ovary, pancreas, and gallbladder)
- Juvenile polyposis syndrome (JPS)
  Juvenile hamartomatous polyps found mainly in colon
  
  Usually diagnosed in adolescence
  
  Median age – 33 years
- Serrated polyposis syndrome
  Serrated polyps usually >10 mm
  
  Risk for CRC not as high as in other syndromes
- Li-Fraumeni syndrome
  Lifetime risk of CRC increased
  
  Core malignancies are sarcoma, as well as breast, adrenocortical, and brain tumors
- Cowden syndrome
  Polyps often >50 with various histologies (eg, hyperplastic, adenomatous, hamartomatous)
  
  Risk much lower for CRC than FAP or MAP
  
  Increased risk for other malignancy (eg, skin, breast, thyroid, endometrial, brain)

Tests generally appear in the order most useful for common clinical situations. Click on number for test-specific information in the ARUP Laboratory Test Directory.

Mismatch Repair by Immunohistochemistry 0049302
Method: Qualitative Immunohistochemistry

Recommended Use

First-line screening test for Lynch syndrome (LS)

Highly recommended prior to ordering MMR germline mutation analysis gene testing; results direct subsequent genetic diagnostic testing

Determines MMR protein expression (MLH1, MSH2, PMS2, and MSH6) in tumor tissue

Refer to Additional Technical Information document for further content

Clinical sensitivity – 90% (NCCN, 2016)

Limitations

~10% of individuals with LS will have IHC tests that show normal staining of the MMR proteins

Because correlation of MSI with IHC is not 100%, direct testing of MSI by PCR may be helpful

Screens for LS only and does not evaluate other hereditary causes of colorectal cancer (CRC)

Occult Blood, Fecal by Immunoassay 2007190
Method: Quantitative Immunoassay

Recommended Use

Screen for CRC

Colonoscopy is the recommended test of choice for individuals >50 years

More sensitive than guaiac testing

Limitations

Less sensitive than colonoscopy

Microsatellite Instability (MSI), HNPCC/Lynch Syndrome, by PCR 0051740
Method: Polymerase Chain Reaction/Fragment Analysis

Recommended Use

First-line screening test for LS

Directs subsequent genetic diagnostic testing for LS

Genes included – MLH1, MSH2, MSH6, PMS2

Refer to Additional Technical Information document for further content

Clinical sensitivity – 90%

Limitations

15% of sporadic CRCs are also MSI-H

Preoperative chemoradiation of rectal cancer may complicate IHC interpretation and/or decrease tumor mass and make MSI testing difficult

Evaluation of pretreatment biopsies will avoid this limitation

Screens for LS only and does not evaluate other hereditary causes of CRC

Epi proColon 2013906
Method: Polymerase Chain Reaction

Recommended Use

Indicated to screen adults of either sex, ≥50 years, defined as average risk for CRC, who have been offered and have a history of not completing CRC screening

Tests that are available and recommended in the USPSTF 2008 CRC screening guidelines should be offered and declined prior to offering the Epi proColon test

Patients with a positive Epi proColon test result should be referred for diagnostic colonoscopy

The Epi proColon test results should be used in combination with physician's assessment and individual risk factors in guiding patient management

For more information, please refer to the Instructions for Use

Follow-up

Test results should be used in combination with physician's assessment and individual risk factors in guiding patient management

Patients with a positive Epi proColon test result should be referred for diagnostic colonoscopy

Colon Cancer Gene Panel, Somatic 2011616
Method: Mass Spectrometry

Recommended Use

Indicated for individuals with metastatic CRC to guide treatment with anti-EGFR monoclonal antibodies (eg, cetuximab and panitumumab)

More cost effective than ordering multiple single gene assays

Detect mutations in

KRAS – codons 12, 13, 61 (MassARRAY also detects codon 146)
BRAF – codon 600
PIK3CA – codons 542, 545, 1047
NRAS – codons 12, 13, 61

Refer to Additional Technical Information document for further content

Clinical sensitivity

KRAS – activating mutation found in ~40% of CRCs
BRAF – activating mutation found in ~10% of CRCs
PIK3CA – oncogenic mutation found 10-20% of CRCs
NRAS – oncogenic mutation found in ~3% of CRCs

Limitations

Limit of detection – ~10% mutant alleles

Oncogenic mutations outside of codons tested will not be detected

KRAS Mutation Detection with Reflex to BRAF Codon 600 Mutation Detection 2001932
Method: Polymerase Chain Reaction/Pyrosequencing

Recommended Use

Determine eligibility for anti-EGFR (cetuximab and panitumumab) therapy in patients with metastatic CRC

Detect mutations in codons 12, 13, 61

Refer to Additional Technical Information document for further content

Reflex pattern – if KRAS mutation is not detected, BRAF codon 600 mutation detection test will be performed

Clinical sensitivity – activating KRAS mutation found in ~40% of CRCs

Limitations

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codons tested will not be detected

A substantial portion of individuals with wild-type KRAS still fail to respond to anti-EGFR agents, implicating downstream mutations

KRAS Mutation Detection 0040248
Method: Polymerase Chain Reaction/Pyrosequencing

Recommended Use

Predicts response to anti-EGFR and MAPK pathway therapies in a variety of malignancies, including CRC

Clinical sensitivity – activating KRAS mutations found in ~40% of CRCs

Limitations

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codons tested will not be detected

A substantial portion of individuals with wild type KRAS still fail to respond to anti-EGFR agents, implicating downstream mutations

BRAF Codon 600 Mutation Detection by Pyrosequencing 2002498
Method: Polymerase Chain Reaction/Pyrosequencing

Recommended Use

Use to detect activating BRAF mutations at codon 600, which can indicate resistance to anti-EGFR therapy in CRC

This test is also used within Lynch syndrome reflex testing pathway (for CRC specimens only)

Clinical sensitivity – activating BRAF found in ~10% of CRC

Limitations

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codon tested will not be detected

BRAF V600E Mutation Detection in Circulating Cell-Free DNA by Digital Droplet PCR 2013921
Method: Polymerase Chain Reaction

Recommended Use

Determine BRAF V600E mutation status in patients with solid tumors to select candidates for targeted therapy with kinase (BRAF and/or MEK) inhibitors

Monitor response to therapy and disease progression in patients carrying BRAF V600E mutation

NRAS Mutation Detection by Pyrosequencing 2003123
Method: Polymerase Chain Reaction/Pyrosequencing

Recommended Use

Predict response to anti-EGFR and MAPK pathway therapies in a variety of malignancies, including CRCs

Detect activating NRAS mutations (codons 12, 13, 61) associated with relative resistance to anti-EGFR therapy

Clinical sensitivity – oncogenic NRAS mutation found in ~3% of CRCs

Limitations

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codons tested are not detected

Presence or absence of mutations does not guarantee a positive response to anti-EGFR therapies

PTEN by Immunohistochemistry 2004115
Method: Immunohistochemistry

Recommended Use

Detect loss of PTEN expression in tumor tissue

Possibly associated with relative resistance to anti-EGFR therapy

Does not include interpretation

Stained and returned to client pathologist; consultation available if needed

Clinical sensitivity/specificity – loss of expression found in up to 50% of CRCs

PTEN with Interpretation by Immunohistochemistry 2007031
Method: Immunohistochemistry

Recommended Use

Detect loss of PTEN expression in tumor tissue

Possibly associated with relative resistance to anti-EGFR therapy

Stained and returned to client pathologist; includes interpretation

Clinical sensitivity/specificity – loss of expression found in up to 50% of CRCs

PIK3CA Mutation Detection (INACTIVE as of 08/21/17: Refer to 2011616) 2004510
Method: Polymerase Chain Reaction/Pyrosequencing

Recommended Use

Predict response to anti-EGFR and AKT/mTOR pathway therapies in a variety of malignancies, including CRCs

Detect activating PIK3CA mutations in exons 9 (codon 542, 545) and 20 (codon 1047)

Clinical sensitivity – oncogenic PIK3CA mutation found in 10-20% of CRCs, mostly exons 9 (60-65%) or 20 (20-25%)

Limitations

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codons tested will not be detected

Presence or absence of mutations does not guarantee a response or lack of response to anti-EGFR therapy

Familial Mutation, Targeted Sequencing 2001961
Method: Polymerase Chain Reaction/Sequencing

Recommended Use

Useful when a pathogenic familial variant identifiable by sequencing is known

Consultation with a genetics counselor is advised

Familial Adenomatous Polyposis Panel: (APC) Sequencing and Deletion/Duplication, (MUTYH) 2 Mutations 2004915
Method: Polymerase Chain Reaction/Sequencing/Multiplex Ligation-dependent Probe Amplification

Recommended Use

Preferred diagnostic or predictive test for familial adenomatous polyposis (FAP) and MUTYH-associated adenomatous polyposis (MAP)

APC gene – up to 90% of pathogenic variants detected by sequencing; ~8-12% of pathogenic variants detected by deletion/duplication testing

MAP – Y165C and G382D represent 85% of pathogenic MUTYH gene variants in Northern European Caucasians

Refer to Additional Technical Information for further content

Clinical sensitivity

Classic FAP – ~95%
Attenuated FAP – <30%
MAP in Northern European Caucasians – 85% (must detect 2 pathogenic variants for diagnosis)

Limitations

APC gene

Deep intronic or regulatory region variants will not be identified
Breakpoints of large deletions/duplications will not be determined

Only 2 pathogenic MUTYH gene variants will be tested – Y165C and G382D

Diagnostic errors can occur due to rare sequence variations

Carcinoembryonic Antigen 0080080
Method: Quantitative Electrochemiluminescent Immunoassay

Recommended Use

Monitor tumor recurrence

Limitations

Not sensitive or specific enough for screening in the general population

MUTYH-Associated Polyposis (MUTYH) 2 Mutations 2004911
Method: Polymerase Chain Reaction/Sequencing

Recommended Use

Acceptable diagnostic or predictive test for MAP in Northern European Caucasians

For non-Caucasians, refer to MUTYH gene sequencing testing

Only 2 pathogenic MUTYH mutations are tested – c.494A>G (p.Y165C) and c.1145G>A (p.G382D)

Limitations

Not detected – large deletions or duplications; deep intronic, regulatory region, or promoter pathogenic mutation

Diagnostic errors can occur due to rare sequence variations

Circulating Tumor Cell Count 0093399
Method: Immunomagnetic Separation/Immunofluorescent Stain/Computer Assisted Analysis

Recommended Use

Use in metastatic tumors in conjunction with clinical data and imaging to monitor response to therapy and disease progression

Cutoffs vary by tumor cell type

Limitations

CTC test is not as accurate as imaging in assessing whether a patient has progressive disease

Doxorubicin therapy patients – allow at least 7 days following administration of dose before testing

Not detected – CTCs that do not express EpCAM; CTCs that express EpCAM but not cytokeratins 8, 18, 19

Serial CTCs should be performed in the same laboratory

MUTYH-Associated Polyposis (MUTYH) 2 Mutations with Reflex to Sequencing 2006307
Method: Polymerase Chain Reaction/Sequencing

Recommended Use

Preferred diagnostic or predictive test for MAP in Caucasians

For non-Caucasians, order MUTYH gene sequencing

Reflex pattern – MUTYH sequencing will be performed if 2 pathogenic mutations are not identified by targeted testing for Y165C and G382D

Gastrointestinal Hereditary Cancer Panel, Sequencing and Deletion/Duplication, 16 Genes 2013449
Method: Massively Parallel Sequencing/Exonic Oligonucleotide-based CGH Microarray/Sequencing/Multiplex Ligation-dependent Probe Amplification

Recommended Use

Confirm a diagnosis of hereditary gastrointestinal (GI) cancer in individuals with a personal or family history of GI cancer and/or polyposis

Includes all 4 MMR genes known to cause LS

Preferred test for individuals with suspected Lynch syndrome or another hereditary GI cancer syndrome

Analysis of specific genes included in this panel may be available individually at ARUP

Genes included – APC, BMPR1A, CDH1, EPCAM, MLH1, MSH2, MSH6, MUTYH, PMS2, PTEN, SDHB, SDHC, SDHD, SMAD4, STK11, TP53

For more information, refer to Additional Technical Information document

Limitations

Diagnostic errors can occur due to rare sequence variations

Not determined or evaluated – Variants in genes not included on the panel; deep intronic and regulatory region variants; breakpoints for large deletions/duplications; sequence changes in EPCAM gene

Deletions/duplications may not be detected in exon 9 in BMPR1A gene; exon 1 in CDH1 and MSH2 genes; exon 8 in PMS2 gene; exons 4, 6, and 7 in STK11 gene

Individuals with hematological malignancy and/or a previous allogenic bone marrow transplant should not undergo molecular genetic testing on peripheral blood specimen; testing of cultured fibroblasts or buccal specimen is required for accurate interpretation of test results; not all predisposing genes are analyzed

Guidelines

American Cancer Society Guidelines for the Early Detection of Cancer. American Cancer Society. Atlanta, GA [Revised Jul 2016; Accessed: Dec 2016]

Choosing Wisely. An initiative of the ABIM Foundation. [Accessed: Jun 2017]

Hampel H, Bennett RL, Buchanan A, Pearlman R, Wiesner GL, Guideline Development Group, American College of Medical Genetics and Genomics Professional Practice and Guidelines Committee and National Society of Genetic Counselors Practice Guidelines Committee. A practice guideline from the American College of Medical Genetics and Genomics and the National Society of Genetic Counselors: referral indications for cancer predisposition assessment. Genet Med. 2015; 17(1): 70-87. PubMed

Lin JS, Piper MA, Perdue LA, Rutter CM, Webber EM, O'Connor E, Smith N, Whitlock EP. Screening for Colorectal Cancer: Updated Evidence Report and Systematic Review for the US Preventive Services Task Force JAMA. 2016; 315(23): 2576-94. PubMed

NCCN Clinical Practice Guidelines in Oncology, Colon Cancer. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Feb 2017]

NCCN Clinical Practice Guidelines in Oncology, Colorectal Cancer Screening. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Feb 2017]

NCCN Clinical Practice Guidelines in Oncology, Genetic/Familial High-Risk Assessment: Colorectal. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Feb 2017]

NCCN Clinical Practice Guidelines in Oncology, Rectal Cancer. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Feb 2017]

Protocol for the Examination of Specimens from Patients with Primary Carcinoma of the Colon and Rectum. Based on AJCC/UICC TNM, 7th ed. Protocol web posting date: Jan 2016. College of American Pathologists (CAP). Northfield, IL [Revised Oct 2013; Accessed: Dec 2016]

Schmoll HJ, Van Cutsem E, Stein A, Valentini V, Glimelius B, Haustermans K, Nordlinger B, van de Velde CJ, Balmana J, Regula J, Nagtegaal ID, Beets-Tan RG, Arnold D, Ciardiello F, Hoff P, Kerr D, Köhne CH, Labianca R, Price T, Scheithauer W, Sobrero A, Tabernero J, Aderka D, Barroso S, Bodoky G, Douillard JY, Ghazaly E, Gallardo J, Garin A, Glynne-Jones R, Jordan K, Meshcheryakov A, Papamichail D, Pfeiffer P, Souglakos I, Turhal S, Cervantes A. ESMO Consensus Guidelines for management of patients with colon and rectal cancer. a personalized approach to clinical decision making. Ann Oncol. 2012; 23(10): 2479-516. PubMed

Sepulveda AR, Hamilton SR, Allegra CJ, Grody W, Cushman-Vokoun AM, Funkhouser WK, Kopetz SE, Lieu C, Lindor NM, Minsky BD, Monzon FA, Sargent DJ, Singh VM, Willis J, Clark J, Colasacco C, Rumble B, Temple-Smolkin R, Ventura CB, Nowak JA. Molecular Biomarkers for the Evaluation of Colorectal Cancer: Guideline From the American Society for Clinical Pathology, College of American Pathologists, Association for Molecular Pathology, and the American Society of Clinical Oncology. J Clin Oncol. 2017; JCO2016719807. PubMed

Stoffel EM, Mangu PB, Gruber SB, Hamilton SR, Kalady MF, Lau MW, Lu KH, Roach N, Limburg PJ, American Society of Clinical Oncology, European Society of Clinical Oncology. Hereditary colorectal cancer syndromes: American Society of Clinical Oncology Clinical Practice Guideline endorsement of the familial risk-colorectal cancer: European Society for Medical Oncology Clinical Practice Guidelines. J Clin Oncol. 2015; 33(2): 209-17. PubMed

Van Cutsem E, Cervantes A, Adam R, Sobrero A, Van Krieken JH, Aderka D, Aguilar A, Bardelli A, Benson A, Bodoky G, Ciardiello F, D'Hoore A, Diaz-Rubio E, Douillard J, Ducreux M, Falcone A, Grothey A, Gruenberger T, Haustermans K, Heinemann V, Hoff P, Köhne C, Labianca R, Laurent-Puig P, Ma B, Maughan T, Muro K, Normanno N, Österlund P, Oyen WJ, Papamichael D, Pentheroudakis G, Pfeiffer P, Price TJ, Punt C, Ricke J, Roth A, Salazar R, Scheithauer W, Schmoll HJ, Tabernero J, Taïeb J, Tejpar S, Wasan H, Yoshino T, Zaanan A, Arnold D. ESMO consensus guidelines for the management of patients with metastatic colorectal cancer. Ann Oncol. 2016; 27(8): 1386-422. PubMed

General References

Bedeir A, Krasinskas AM. Molecular diagnostics of colorectal cancer. Arch Pathol Lab Med. 2011; 135(5): 578-87. PubMed

Chua W, Moore MM, Charles KA, Clarke SJ. Predictive biomarkers of clinical response to targeted antibodies in colorectal cancer. Curr Opin Mol Ther. 2009; 11(6): 611-22. PubMed

Colle R, Cohen R, Cochereau D, Duval A, Lascols O, Lopez-Trabada D, Afchain P, Trouilloud I, Parc Y, Lefevre JH, Fléjou J, Svrcek M, André T. Immunotherapy and patients treated for cancer with microsatellite instability. Bull Cancer. 2017; 104(1): 42-51. PubMed

Cunningham D, Atkin W, Lenz H, Lynch HT, Minsky B, Nordlinger B, Starling N. Colorectal cancer. Lancet. 2010; 375(9719): 1030-47. PubMed

deVos T, Tetzner R, Model F, Weiss G, Schuster M, Distler J, Steiger KV, Grützmann R, Pilarsky C, Habermann JK, Fleshner PR, Oubre BM, Day R, Sledziewski AZ, Lofton-Day C. Circulating methylated SEPT9 DNA in plasma is a biomarker for colorectal cancer. Clin Chem. 2009; 55(7): 1337-46. PubMed

Evaluation of Genomic Applications in Practice and Prevention (EGAPP) Working Group. Recommendations from the EGAPP Working Group: can UGT1A1 genotyping reduce morbidity and mortality in patients with metastatic colorectal cancer treated with irinotecan? Genet Med. 2009; 11(1): 15-20. PubMed

Fearon ER. Molecular genetics of colorectal cancer. Annu Rev Pathol. 2011; 6: 479-507. PubMed

Impact of primary (1º) tumor location on overall survival (OS) and progression-free survival (PFS) in patients (pts) with metastatic colorectal cancer (mCRC): Analysis of CALGB/SWOG 80405 (Alliance). J Clin Oncol. 34, 2016 (suppl; abstr 3504) [Accessed: Dec 2016]

Newton KF, Newman W, Hill J. Review of biomarkers in colorectal cancer. Colorectal Dis. 2012; 14(1): 3-17. PubMed

Ross JS. Clinical implementation of KRAS testing in metastatic colorectal carcinoma: the pathologist's perspective. Arch Pathol Lab Med. 2012; 136(10): 1298-307. PubMed

Sharma SG, Gulley ML. BRAF mutation testing in colorectal cancer. Arch Pathol Lab Med. 2010; 134(8): 1225-8. PubMed

Song L, Li Y. SEPT9: A Specific Circulating Biomarker for Colorectal Cancer. Adv Clin Chem. 2015; 72: 171-204. PubMed

Venook AP. Metastatic Colorectal Cancer: Lessons Learned, Future Possibilities J Natl Compr Canc Netw. 2016; 14(5 Suppl): 666-8. PubMed

Wilkins T, Reynolds PL. Colorectal cancer: a summary of the evidence for screening and prevention. Am Fam Physician. 2008; 78(12): 1385-92. PubMed

Yurgelun MB, Goel A, Hornick JL, Sen A, Turgeon DK, Ruffin MT, Marcon NE, Baron JA, Bresalier RS, Syngal S, Brenner DE, Boland R, Stoffel EM. Microsatellite instability and DNA mismatch repair protein deficiency in Lynch syndrome colorectal polyps. Cancer Prev Res (Phila). 2012; 5(4): 574-82. PubMed

References from the ARUP Institute for Clinical and Experimental Pathology®

Affolter K, Samowitz W, Tripp S, Bronner MP. BRAF V600E mutation detection by immunohistochemistry in colorectal carcinoma. Genes Chromosomes Cancer. 2013; 52(8): 748-52. PubMed

Burt RW, Leppert MF, Slattery ML, Samowitz WS, Spirio LN, Kerber RA, Kuwada SK, Neklason DW, Disario JA, Lyon E, Hughes P, Chey WY, White RL. Genetic testing and phenotype in a large kindred with attenuated familial adenomatous polyposis. Gastroenterology. 2004; 127(2): 444-51. PubMed

Campbell PT, Curtin K, Ulrich CM, Samowitz WS, Bigler J, Velicer CM, Caan B, Potter JD, Slattery ML. Mismatch repair polymorphisms and risk of colon cancer, tumour microsatellite instability and interactions with lifestyle factors. Gut. 2009; 58(5): 661-7. PubMed

Curtin K, Samowitz WS, Wolff RK, Caan BJ, Ulrich CM, Potter JD, Slattery ML. MSH6 G39E polymorphism and CpG island methylator phenotype in colon cancer. Mol Carcinog. 2009; 48(11): 989-94. PubMed

Curtin K, Samowitz WS, Wolff RK, Ulrich CM, Caan BJ, Potter JD, Slattery ML. Assessing tumor mutations to gain insight into base excision repair sequence polymorphisms and smoking in colon cancer. Cancer Epidemiol Biomarkers Prev. 2009; 18(12): 3384-8. PubMed

Curtin K, Slattery ML, Ulrich CM, Bigler J, Levin TR, Wolff RK, Albertsen H, Potter JD, Samowitz WS. Genetic polymorphisms in one-carbon metabolism: associations with CpG island methylator phenotype (CIMP) in colon cancer and the modifying effects of diet. Carcinogenesis. 2007; 28(8): 1672-9. PubMed

Curtin K, Ulrich CM, Samowitz WS, Bigler J, Caan B, Potter JD, Slattery ML. Thymidylate synthase polymorphisms and colon cancer: associations with tumor stage, tumor characteristics and survival. Int J Cancer. 2007; 120(10): 2226-32. PubMed

Eliason K, Hendrickson BC, Judkins T, Norton M, Leclair B, Lyon E, Ward B, Noll W, Scholl T. The potential for increased clinical sensitivity in genetic testing for polyposis colorectal cancer through the analysis of MYH mutations in North American patients. J Med Genet. 2005; 42(1): 95-6. PubMed

Ferrández A, Pho L, Solomon C, Samowitz WS, Kuwada SK, Knecht TP, Gilfeather M, Burt RW. An evidence-based, multidisciplinary approach to the clinical considerations, management, and surveillance of adrenal lesions in familial adenomatous polyposis: report of three cases. Dis Colon Rectum. 2006; 49(11): 1781-90. PubMed

Neklason DW, Thorpe BL, Ferrández A, Tumbapura A, Boucher K, Garibotti G, Kerber RA, Solomon CH, Samowitz WS, Fang JC, Mineau GP, Leppert MF, Burt RW, Kuwada SK. Colonic adenoma risk in familial colorectal cancer--a study of six extended kindreds. Am J Gastroenterol. 2008; 103(10): 2577-84. PubMed

Patil DT, Bronner MP, Portier BP, Fraser CR, Plesec TP, Liu X. A five-marker panel in a multiplex PCR accurately detects microsatellite instability-high colorectal tumors without control DNA. Diagn Mol Pathol. 2012; 21(3): 127-33. PubMed

Pellatt DF, Stevens JR, Wolff RK, Mullany LE, Herrick JS, Samowitz W, Slattery ML. Expression Profiles of miRNA Subsets Distinguish Human Colorectal Carcinoma and Normal Colonic Mucosa. Clin Transl Gastroenterol. 2016; 7: e152. PubMed

Rowe LR, Bentz BG, Bentz JS. Detection of BRAF V600E activating mutation in papillary thyroid carcinoma using PCR with allele-specific fluorescent probe melting curve analysis. J Clin Pathol. 2007; 60(11): 1211-5. PubMed

Salk JJ, Bansal A, Lai LA, Crispin DA, Ussakli CH, Horwitz MS, Bronner MP, Brentnall TA, Loeb LA, Rabinovitch PS, Risques RA. Clonal expansions and short telomeres are associated with neoplasia in early-onset, but not late-onset, ulcerative colitis. Inflamm Bowel Dis. 2013; 19(12): 2593-602. PubMed

Samadder J, Neklason DW, Boucher KM, Byrne KR, Kanth P, Samowitz W, Jones D, Tavtigian SV, Done MW, Berry T, Jasperson K, Pappas L, Smith L, Sample D, Davis R, Topham MK, Lynch P, Strait E, McKinnon W, Burt RW, Kuwada SK. Effect of Sulindac and Erlotinib vs Placebo on Duodenal Neoplasia in Familial Adenomatous Polyposis: A Randomized Clinical Trial. JAMA. 2016; 315(12): 1266-75. PubMed

Samowitz WS, Albertsen H, Sweeney C, Herrick J, Caan BJ, Anderson KE, Wolff RK, Slattery ML. Association of smoking, CpG island methylator phenotype, and V600E BRAF mutations in colon cancer. J Natl Cancer Inst. 2006; 98(23): 1731-8. PubMed

Samowitz WS, Broaddus R, Iacopetta B, Goldblatt J. PCR versus immunohistochemistry for microsatellite instability. J Mol Diagn. 2008; 10(2): 181-2; author reply 181. PubMed

Samowitz WS, Curtin K, Wolff RK, Albertsen H, Sweeney C, Caan BJ, Ulrich CM, Potter JD, Slattery ML. The MLH1 -93 G>A promoter polymorphism and genetic and epigenetic alterations in colon cancer. Genes Chromosomes Cancer. 2008; 47(10): 835-44. PubMed

Samowitz WS, Curtin K, Wolff RK, Tripp SR, Caan BJ, Slattery ML. Microsatellite instability and survival in rectal cancer. Cancer Causes Control. 2009; 20(9): 1763-8. PubMed

Samowitz WS, Ogino S. DNA methylation in breast and colorectal cancers. Mod Pathol. 2008; 21(8): 1054; author reply 1054-5. PubMed

Samowitz WS, Slattery ML, Sweeney C, Herrick J, Wolff RK, Albertsen H. APC mutations and other genetic and epigenetic changes in colon cancer. Mol Cancer Res. 2007; 5(2): 165-70. PubMed

Samowitz WS, Wolff RK, Curtin K, Sweeney C, Ma K, Andersen K, Levin TR, Slattery ML. Interactions between CYP2C9 and UGT1A6 polymorphisms and nonsteroidal anti-inflammatory drugs in colorectal cancer prevention. Clin Gastroenterol Hepatol. 2006; 4(7): 894-901. PubMed

Samowitz WS, Wolff RK, Ma KN, Andersen K, Caan B, Slattery ML. Polymorphisms in insulin-related genes predispose to specific KRAS2 and TP53 mutations in colon cancer. Mutat Res. 2006; 595(1-2): 117-24. PubMed

Samowitz WS. Genetic and epigenetic changes in colon cancer. Exp Mol Pathol. 2008; 85(1): 64-7. PubMed

Samowitz WS. The CpG island methylator phenotype in colorectal cancer. J Mol Diagn. 2007; 9(3): 281-3. PubMed

Shaco-Levy R, Jasperson KW, Martin K, Samadder J, Burt RW, Ying J, Bronner MP. Morphologic characterization of hamartomatous gastrointestinal polyps in Cowden syndrome, Peutz-Jeghers syndrome, and juvenile polyposis syndrome. Hum Pathol. 2016; 49: 39-48. PubMed

Slattery ML, Curtin K, Sweeney C, Levin TR, Potter J, Wolff RK, Albertsen H, Samowitz WS. Diet and lifestyle factor associations with CpG island methylator phenotype and BRAF mutations in colon cancer. Int J Cancer. 2007; 120(3): 656-63. PubMed

Slattery ML, Curtin K, Wolff R, Ma KN, Sweeney C, Murtaugh M, Potter JD, Levin TR, Samowitz W. PPARgamma and colon and rectal cancer: associations with specific tumor mutations, aspirin, ibuprofen and insulin-related genes (United States). Cancer Causes Control. 2006; 17(3): 239-49. PubMed

Slattery ML, Curtin K, Wolff RK, Boucher KM, Sweeney C, Edwards S, Caan BJ, Samowitz W. A comparison of colon and rectal somatic DNA alterations. Dis Colon Rectum. 2009; 52(7): 1304-11. PubMed

Slattery ML, Herrick JS, Mullany LE, Valeri N, Stevens J, Caan BJ, Samowitz W, Wolff RK. An evaluation and replication of miRNAs with disease stage and colorectal cancer-specific mortality Int J Cancer. 2015; 137(2): 428-38. PubMed

Slattery ML, Herrick JS, Pellatt DF, Stevens JR, Mullany LE, Wolff E, Hoffman MD, Samowitz WS, Wolff RK. MicroRNA profiles in colorectal carcinomas, adenomas and normal colonic mucosa: variations in miRNA expression and disease progression. Carcinogenesis. 2016; 37(3): 245-61. PubMed

Slattery ML, Wolff RK, Curtin K, Fitzpatrick F, Herrick J, Potter JD, Caan BJ, Samowitz WS. Colon tumor mutations and epigenetic changes associated with genetic polymorphism: insight into disease pathways. Mutat Res. 2009; 660(1-2): 12-21. PubMed

Sweeney C, Boucher KM, Samowitz WS, Wolff RK, Albertsen H, Curtin K, Caan BJ, Slattery ML. Oncogenetic tree model of somatic mutations and DNA methylation in colon tumors. Genes Chromosomes Cancer. 2009; 48(1): 1-9. PubMed

Szankasi P, Reading S, Vaughn CP, Prchal JT, Bahler DW, Kelley TW. A quantitative allele-specific PCR test for the BRAF V600E mutation using a single heterozygous control plasmid for quantitation: a model for qPCR testing without standard curves. J Mol Diagn. 2013; 15(2): 248-54. PubMed

Tomsic J, Senter L, Liyanarachchi S, Clendenning M, Vaughn CP, Jenkins MA, Hopper JL, Young J, Samowitz W, de la Chapelle A. Recurrent and founder mutations in the PMS2 gene. Clin Genet. 2013; 83(3): 238-43. PubMed

Vaughn CP, Baker CL, Samowitz WS, Swensen JJ. The frequency of previously undetectable deletions involving 3' Exons of the PMS2 gene. Genes Chromosomes Cancer. 2013; 52(1): 107-12. PubMed

Vaughn CP, Lyon E, Samowitz WS. Confirmation of single exon deletions in MLH1 and MSH2 using quantitative polymerase chain reaction. J Mol Diagn. 2008; 10(4): 355-60. PubMed

Walter AW, Ennis S, Best H, Vaughn CP, Swensen JJ, Openshaw A, Gripp KW. Constitutional mismatch repair deficiency presenting in childhood as three simultaneous malignancies. Pediatr Blood Cancer. 2013; 60(11): E135-6. PubMed