• Assess need to test individuals or their family members for hereditary nonpolyposis colorectal cancer (HNPCC)-associated mutations

HNPCC, an autosomal dominant condition caused by mutations in mismatch repair (MMR) genes, accounts for about 3% to 5% of colorectal cancers. In individuals with an MMR mutation, the risk of developing colorectal cancer is ~80% by age 75;¹ these mutations may also predispose to endometrial, ovarian, stomach, and other forms of cancer. Although HNPCC may have a somewhat better prognosis than other forms of colorectal cancer, it is characterized by rapid progression from adenomatous polyps to malignant lesions. Identifying HNPCC in affected individuals is important, as close surveillance of at-risk family members has been found to reduce the rate of colorectal cancer and overall mortality by >60%.² Family members with an MMR mutation require aggressive follow-up (colonoscopy every 1–2 years), while those without the familial mutation are at the same risk as the general population and do not require intensive screening.

HNPCC can be diagnosed on the basis of clinical and family history,³ as well as by direct detection of MMR mutations. In suspected HNPCC patients without a known familial mutation, the first step in the genetic diagnosis is identifying microsatellite instability (MSI) in tumor cells. Microsatellites are sequences of repetitive DNA with repeating units of 1 to 7 base pairs. These regions are prone to replication errors, which are normally repaired by MMR enzymes. If one of the MMR enzymes is impaired, the microsatellites become unstable and will have varying repeat numbers at specific loci. Although MSI does not seem to have any clinical effect, it is a marker of faulty DNA repair; cancer arises when faulty DNA repair leads to mutations in tumor suppressor or oncogenes. MSI is found in most HNPCCs but only about 15% of sporadic colorectal tumors,^4,5 and therefore cannot be used alone for diagnosis of HNPCC. However, a finding of MSI strongly indicates the presence of MMR mutations and the need for further genetic testing.⁵

The National Cancer Institute has recommended a panel of 5 microsatellites for MSI screening: BAT 25 and BAT 26 (mononucleotide repeats), D2S123, D5S346, and D17S250 (dinucleotide repeats).⁶

• Individuals who meet any of the Bethesda criteria (Table) and in whom a familial mutation is not known

Submit 1 formalin-fixed, paraffin-embedded tissue block with representative tumor plus 5 mL whole blood in a lavender-top (EDTA) tube or yellow-top (ACD) tube (2 mL minimum). Ship at room temperature; do not freeze.

• Paraffin sections containing ≥30% tumor cells are processed with selective scraping of the tumor-rich area; paraffin sections containing <30% tumor cells are processed with laser capture microdissection

• Multiplex polymerase chain reaction (PCR) amplification of 5 STR loci (BAT 25, BAT 26, D2S123, D5S346, D17S250) from tumor and normal tissue

• Separation of amplification products by capillary electrophoresis

• Data analysis with GeneMapper™ software detects peaks and determines allele size. The tumor and normal peaks are visually compared to detect MSI.

• Results reported as MSI-high (MSI-H) if ≥2 loci are positive; MSI-low (MSI-L) if 1 locus is positive; microsatellite stable if all loci are negative

• Aliases: HNPCC; Lynch syndrome; replication error (RER+)

• CPT codes*: 83890 x2, 83907, 83900, 83901 x3, 83900, 83901 x3, 83909 x2, 83912

MSI not detected

Individuals with MSI-H tumors should be screened for germline mutations in MLH1 and MSH2, and in MSH6 if no MLH1 or MSH2 mutations are found. Although most patients with HNPCC exhibit MSI-H, up to 10% of cases exhibit MSI-L or negative results. Thus, MMR mutation analysis should be considered if a strong suspicion of HNPCC exists; indeed MSI is frequently absent in tumors associated with MSH6 mutations.⁸