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Test ID: MLHPB MLH1 Hypermethylation Analysis, Blood

Useful For

As an adjunct to positive hypermethylation in tumor to distinguish between somatic and germline hypermethylation


As an adjunct to negative MLH1 germline testing in cases where colon or endometrial tumor demonstrates microsatellite instability-H (MSI-H) and loss of MLH1 protein expression

Method Name

Polymerase Chain Reaction (PCR)

Reporting Name

MLH1 Hypermethylation Analys, Blood

Specimen Type


Shipping Instructions

Specimen preferred to arrive within 96 hours of draw.

Specimen Required

Patient Preparation: A previous bone marrow transplant from an allogenic donor will interfere with testing. Call 800-533-1710 for instructions for testing patients who have received a bone marrow transplant.


Preferred: Lavender top (EDTA) or yellow top (ACD)

Acceptable: Any anticoagulant

Specimen Volume: 3 mL

Collection Instructions:

1. Invert several times to mix blood.

2. Send specimen in original tube.

Specimen Minimum Volume

1 mL

Specimen Stability Information

Specimen Type Temperature Time Special Container
Varies Ambient (preferred)

Clinical Information

Lynch syndrome/hereditary nonpolyposis colorectal cancer (HNPCC) is an autosomal dominant hereditary cancer syndrome associated with germline mutations in the mismatch repair genes, MLH1, MSH2, MSH6, and PMS2. Deletions within the 3-prime end of the EPCAM gene have also been associated with Lynch syndrome/HNPCC, as this leads to inactivation of the MSH2 promoter. 


Lynch syndrome/HNPCC is predominantly characterized by significantly increased risks for colorectal and endometrial cancer. The lifetime risk for colorectal cancer is highly variable and dependent on the gene involved. The risk for colorectal cancer associated MLH1 and MSH2 mutations (approximately 50%-80%) is generally higher than the risks associated with mutations in the other Lynch syndrome/HNPCC-related genes and the lifetime risk for endometrial cancer (approximately 25%-60%) is also highly variable. Other malignancies within the tumor spectrum include gastric cancer, ovarian cancer, hepatobiliary and urinary tract carcinomas, and small bowel cancer. The lifetime risks for these cancers are <15%. Of the 4 mismatch repair genes, mutations within the PMS2 gene confer the lowest risk for any of the tumors within the Lynch syndrome/HNPCC spectrum.


Several clinical variants of Lynch syndrome/HNPCC have been defined. These include Turcot syndrome, Muir-Torre syndrome, and homozygous mismatch repair mutations (also called constitutional mismatch repair deficiency syndrome). Turcot syndrome and Muir-Torre syndrome are associated with increased risks for cancers within the tumor spectrum described, but also include brain and central nervous system malignancies and sebaceous carcinomas, respectively. Homozygous mismatch repair mutations, characterized by the presence of biallelic deleterious mutations within a mismatch repair gene, are associated with a different clinical phenotype defined by hematologic and brain cancers, cafe au lait macules, and childhood colon or small bowel cancer.


There are several strategies for evaluating individuals whose personal or family history of cancer is suggestive of Lynch syndrome/HNPCC. One such strategy involves testing the tumors from suspected individuals for microsatellite instability (MSI) and/or immunohistochemistry (IHC) for the presence or absence of defective DNA mismatch repair. It is important to note, however, that the MSI-H tumor phenotype is not restricted to inherited cancer cases; approximately 20% of sporadic colon cancers are MSI-H. Thus, MSI-H does not distinguish between a somatic (sporadic) and a germline (inherited) mutation, nor does it identify which gene is involved. Although IHC analysis is helpful in identifying the responsible gene, it also does not distinguish between somatic and germline defects.


Defective mismatch repair in sporadic colon cancer is most often due to an abnormality in MLH1, and the most common cause of gene inactivation is promoter hypermethylation (epigenetic silencing). A specific mutation in the BRAF gene (V600E) has been shown to be present in approximately 70% of tumors with hypermethylation of the MLH1 promoter. Importantly, the V600E mutation is rarely identified in cases with germline MLH1 mutations. Thus, direct assessment of MLH1 promoter methylation status and testing for the BRAF V600E mutation can be used to help distinguish between a germline mutation and epigenetic/somatic inactivation of MLH1. Tumors that have the BRAF V600E mutation and demonstrate MLH1 promoter hypermethylation are almost certainly sporadic, whereas tumors that show neither are most often caused by an inherited mutation.


However, individuals with tumor hypermethylation may additionally have MLH1 promoter hypermethylation consistent with germline inactivation. Individuals with germline inactivation of MLH1 by promoter hypermethylation are at an increased risk for Lynch syndrome/HNPCC-related tumors. In contrast to sequence mutations in MLH1, current evidence suggests that the risk of transmitting germline MLH1 promoter hypermethylation is <50%.

Reference Values

Interpretive report will be provided.


An interpretive report will be provided.

Clinical Reference

1. Hitchins MP, Ward RL: Constitutional (germline) MLH1 epimutation as an aetiological mechanism for hereditary non-polyposis colorectal cancer. J Med Genet 2009;46(12):793-802

2. Hitchins M, Williams R, Cheong K, et al: MLH1 germline epimutations as a factor in hereditary nonpolyposis colorectal cancer. Gastroenterology 2005;129(5):1392-1399

3. Niessen RC, Hofstra RM, Westers H, et al: Germline hypermethylation of MLH1 and EPCAM deletions are a frequent cause of Lynch syndrome. Genes Chromosomes Cancer 2009;48(8):737-744

4. Valle L, Carbonell P, Fernandez V, et al: MLH1 germline epimutations in selected patients with early-onset non-polyposis colorectal cancer. Clin Genet 2007;71(3):232-237

Day(s) Performed


Report Available

8 to 12 days

Test Classification

This test was developed, and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the US Food and Drug Administration.

CPT Code Information


LOINC Code Information

Test ID Test Order Name Order LOINC Value
MLHPB MLH1 Hypermethylation Analys, Blood 97760-3


Result ID Test Result Name Result LOINC Value
52906 Result Summary 50397-9
52907 Result 82939-0
52908 Interpretation 69047-9
52909 Reason for Referral 42349-1
52910 Specimen 31208-2
52911 Source 31208-2
52912 Released By 18771-6

Testing Algorithm

See Lynch Syndrome Testing Algorithm

Mayo Clinic Laboratories | Genetics and Pharmacogenomics Catalog Additional Information: