Lynch Syndrome - Hereditary Nonpolyposis Colorectal Cancer (HNPCC)

Primary Author: Samowitz, Wade S., MD.

  • Key Points
  • Diagnosis
  • Algorithms
  • Screening
  • Monitoring
  • Background
  • Lab Tests
  • References
  • Related Topics
  • Videos

Lynch syndrome (LS) is an autosomal dominant inherited cancer syndrome that predisposes an individual to colorectal, endometrial, gastric, ovarian, upper urinary tract, and other cancers. Several organizations recommend universal screening of all colorectal cancer (CRC) specimens (AGA, 2015; ASCO, 2014; ESMO, 2013; NCCN, 2016) and all endometrial cancer patients (up to age 50, NCCN, 2016) for LS. In most situations, it is most effective to first evaluate suspected LS with immunohistochemistry (IHC) or polymerase chain reaction (PCR), as only 2-4% of CRCs are LS (NCCN, 2016). However, if strong suspicion exists (eg, family history, cancer at a young age), it is reasonable to proceed to single or full-panel genetic testing.

Mismatch repair (MMR) deficiency

  • Presence of MMR deficiency helps identify patients who may have LS
    • MMR deficiency also occurs in ~15% of sporadic CRC .
  • Definitive diagnosis of LS requires differentiating colorectal tumors with MMR deficiency due to a sporadic somatic event from colorectal tumors with MMR deficiency due to a LS germline mutation
  • MMR genes where LS-causing mutations can occur
    • MLH1
    • MSH2
    • MSH6
    • PMS2
    • EPCAM deletions causing methylation of MSH2

Microsatellite instability (MSI)

  • Functions as a surrogate marker for MMR deficiency
    • MSI is caused by the loss of MMR activity

Screening and diagnostic testing for LS

Screening and Diagnostic Testing

Initial testing*

Typically, IHC or PCR testing is used initially to eliminate expense of full gene sequencing for the vast majority of tumors that lack MMR deficiency. Both tests are sensitive and usually produce concordant results. However, it is reasonable to proceed to germline multigene analysis instead if strong suspicion of LS exists (eg, family history, cancer at a young age).

Test

Test Interpretation

IHC  

  • Involves staining of tumor tissue for protein expression of 4 MMR genes known to be mutated in LS (MLH1, MSH2, MSH6, and PMS2)
  • Pattern of protein loss identified on IHC directs mutational testing
  • 5-10% false-negative rate (NCCN, 2016)
  • Isolated PMS2 loss has been associated with germline MLH1 mutation

IHC Result

Likely Gene Mutation

MLH1, PMS2 loss

MLH1

MSH2, MSH6 loss

MSH2

MSH6

MSH6

PMS2

PMS2

BRAF V600E or MLH1 methylation

  • When MLH1 loss is identified using IHC, perform prior to LS mutation analysis
  • Loss of MLH1 is either due to acquired hypermethylation in sporadic tumors or a germline mutation in LS
    • BRAF mutations uncommon in LS but common in sporadic CRC of MSI pathway
  • NOTE: BRAF testing is not appropriate for endometrial cancer; use only methylation testing

If BRAF or MLH1 hypermethylation is positive

  • LS unlikely; probably sporadic colorectal cancer
  • LS gene mutation analysis not necessary unless high suspicion for LS still exists

PCR (panel of five mononucleotide microsatellites)

  • Detects expansion or contraction of microsatellite repeats in the tumor
  • Does not detect which specific MMR protein is deficient
  • Patient tumor tissue is compared to normal tissue
  • Highest sensitivity achieved when PCR is combined with IHC
  • Useful for MSI testing when IHC testing is negative despite high clinical suspicion of LS

Determination of MSI

High – 2 or more markers with instability

  • Consider gene mutation testing
  • IHC can be used to target specific mutation testing

Indeterminate – 1 marker with instability

  • Instability in even 1 mononucleotide repeat can be associated with LS – suggest IHC

Stable – no markers detected with instability

  • Very low risk of LS
  • Add IHC and possibly gene mutation testing if high suspicion of LS exists

Direct gene analysis (germline multi-gene analysis panel or single gene analysis available)

Consider single gene testing when

  • IHC result indicates gene to target
  • Mutation is known

Consider germline multigene panel if strong suspicion exists

  • Family history of hereditary cancer syndrome
  • Cancer at a young age

Mutational testing of MMR genes is gold standard for diagnosing LS

*Refer to Lynch Syndrome Testing (HNPCC) algorithm

Based on NCCN, 2016 and AGA, 2015; US Multi-Society Task Force on Colorectal Cancer, 2014

Indications for Testing

Laboratory Testing

  • For additional information regarding testing strategies, refer to the following

Prognosis

  • Patients with MMR-deficient CRC have improved prognosis compared to stage-matched patients without MMR deficiency
  • Refer to Key Points section for discussion of screening tests
  • For further discussion of Lynch syndrome screening, please refer to the following references
    • NCCN guidelines, Genetic/Familial High Risk Assessment (2016)
    • ASCO (2015)
    • ESMO (2013)
    • USPSTF (2016)
  • Recommend frequent monitoring for patients diagnosed with Lynch syndrome (LS)
  • For further discussion of LS monitoring, please refer to NCCN guidelines, Genetic/Familial High Risk 

Colorectal cancer (CRC) exhibits the characteristics of familial clustering in ~10-15% of cases. The most common cause of hereditary CRC is Lynch syndrome (LS), also known as hereditary nonpolyposis colorectal cancer (HNPCC). LS is caused by a germline mutation in one of the genes within the DNA mismatch repair (MMR) system.

Epidemiology

  • Prevalence
    • 1/440 individuals in general population (AGA, 2015)
    • 2-4% of CRC (NCCN, 2016)
  • Age at presentation – mutation dependent (44-66 years mean)
    • >75% risk of developing CRC by 70 years
  • Sex – M:F, equal

Inheritance

  • Autosomal dominant with incomplete penetrance
  • Germline mutations in 1 of 4 DNA MMR genes
    • MLH1
    • MSH2
      • Small percentage of MSH2 inactivation is due to EPCAM deletions (included with MSH2 testing)
    • MSH6
    • PMS2
    • Inheritance of homozygous mutations for any of the above is termed biallelic MMR deficiency (BMMR-D)
  • MMR gene mutations

MMR Gene Mutations

Mutations include point mutations and large genomic deletions or rearrangements

Common mutations

~90% of mutations in LS

MLH1 (mutL homolog1)

MSH2 (mutS homolog 2)

Less common mutations

MSH6 (mutS homolog 6)

PMS2 (postmeiotic segregation increased 2)

Heterodimeric complexes

Obligatory partners

  • MLH1 and PMS2
    • MLH1 mutation usually leads to concomitant PMS2 loss due to PMS2 degradation
  • MSH2 and MSH6
    • MSH2 mutation usually leads to concomitant MSH6 loss due to MSH6 degradation
  • MSH6 or PMS2 losses or mutations are usually not associated with any other loss

Clinical Presentation

  • Early onset of proximal (right side) CRC – often <50 years
    • Multiple metachronous and synchronous tumors are common
  • Early-onset extra colonic tumors – risk depends on mutation present
    • Endometrial
      • Some patients may present with endometrial tumor rather than colon tumor; some MSH6 germline-mutated families present with mainly endometrial tumors
    • Ovarian
    • Small bowel
    • Brain
    • Pancreatic
    • Gastric
    • Renal pelvis
    • Ureter
    • Hepatobiliary tract
  • Penetrance – variable; patients may present with tumors at older age
    • Especially true for MSH6 and PMS2
    • Universal screening will identify these tumors and patients will not be mistakenly classified as sporadic CRC
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 with Reflex to BRAF Codon 600 Mutation and MLH1 Promoter Methylation 2002327
Method: Qualitative Immunohistochemistry/Qualitative Real-time Polymerase Chain Reaction

Recommended Use 

Preferred screening test for Lynch syndrome (LS) in individuals with colorectal cancer (CRC)

Reflex pattern – if MLH1 IHC is abnormal, evaluations of BRAF codon 600 and, possibly, MLH1 methylation are performed

Clinical sensitivity – 90%

Do not use in endometrial cancer

Limitations 

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

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

Follow-up 

Definitive diagnosis of LS requires additional targeted MMR germline mutation studies

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

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

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

For more information, refer to Additional Technical Information document

Mismatch Repair by Immunohistochemistry with Reflex to MLH1 Promoter Methylation 2005270
Method: Qualitative Immunohistochemistry/Qualitative Real-time Polymerase Chain Reaction

Recommended Use 

Preferred reflex screening test for LS in non-CRC tumors (eg, endometrial carcinoma) 

Reflex pattern – if MLH1 IHC is abnormal, MLH1 methylation is performed

Clinical sensitivity – 90%

Limitations 

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

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

Mismatch Repair by Immunohistochemistry 0049302
Method: Qualitative Immunohistochemistry

Recommended Use 

First-line screening test for LS; directs additional molecular diagnostic testing for LS

Highly recommended prior to ordering MMR germline mutation analysis – results guide subsequent genetic testing

Clinical sensitivity – 90%

Limitations 

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

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

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

Recommended Use 

First-line screening test for LS; directs additional molecular diagnostic testing for LS

Clinical sensitivity – 90%

Analytic sensitivity/specificity – >99%

Limitations 

15% of sporadic CRCs are also MSI-H

Preoperative chemoradiation of rectal cancer

  • May complicate IHC interpretation and/or decrease tumor mass
  • May make MSI testing difficult

Evaluation of pretreatment biopsies will avoid this limitation

HNPCC/Lynch Syndrome (MLH1) Sequencing and Deletion/Duplication 0051650
Method: Polymerase Chain Reaction/Sequencing/Multiplex Ligation-dependent Probe Amplification

Recommended Use 

Detect germline MLH1 mutations

Use in MMR-deficient carcinoma with suggestive IHC (loss of MLH1 and PMS2 protein), absence of BRAF codon 600 mutation, and normal MLH1 methylation studies

Clinical sensitivity – >95% (combined sensitivity across all 4 genes [MLH1, MSH2, MSH6, and PMS2])

Analytic sensitivity/specificity – 99%

Limitations 

Diagnostic errors can occur due to rare sequence variations

Breakpoints of large deletions/duplications will not be determined

Regulatory region mutations, deep intronic mutations, and mutations in genes other than MLH1 will not be detected 

HNPCC/Lynch Syndrome (MSH2) Sequencing and Deletion/Duplication 0051654
Method: Polymerase Chain Reaction/Sequencing/Multiplex Ligation-dependent Probe Amplification

Recommended Use 

Detect germline MSH2 mutations

Use in MMR-deficient carcinoma with suggestive IHC (loss of MSH2 and MSH6 proteins)

Detects large MSH2 deletions and EPCAM 3 prime deletions

Clinical sensitivity – >95% (combined sensitivity across all 4 genes [MLH1, MSH2, MSH6, and PMS2])

Analytic sensitivity/specificity – 99%

Limitations 

Diagnostic errors can occur due to rare sequence variations

Regulatory region mutations, deep intronic mutations, and mutations in genes other than MSH2 will not be detected 

HNPCC/Lynch Syndrome (MSH6) Sequencing and Deletion/Duplication 0051656
Method: Polymerase Chain Reaction/Sequencing/Multiplex Ligation-dependent Probe Amplification

Recommended Use 

Detect germline MSH6 mutations

Use in MMR-deficient carcinoma with suggestive IHC (isolated loss of MSH6 protein)

Clinical sensitivity – >95% (combined sensitivity across all 4 genes [MLH1, MSH2, MSH6, and PMS2])

Analytic sensitivity/specificity – 99%

Limitations 

Diagnostic errors can occur due to rare sequence variations

Regulatory region mutations, deep intronic mutations, and mutations in genes other than MSH6 will not be detected 

HNPCC/Lynch Syndrome (PMS2) Sequencing and Deletion/Duplication 0051737
Method: Polymerase Chain Reaction/Sequencing/Multiplex Ligation-dependent Probe Amplification

Recommended Use 

Detect germline PMS2 mutations

Use in MMR-deficient carcinoma with suggestive IHC (isolated loss of PMS2 protein)

Clinical sensitivity – >95% (combined sensitivity across all 4 genes [MLH1, MSH2, MSH6, and PMS2])

Analytic sensitivity/specificity – 99%

Limitations 

Diagnostic errors can occur due to rare sequence variations

Regulatory region mutations, deep intronic mutations, and mutations in genes other than PMS2 will not be detected 

BRAF Codon 600 Mutation Detection with Reflex to MLH1 Promoter Methylation 0051750
Method: Polymerase Chain Reaction/Pyrosequencing

Recommended Use 

Recommended reflex test for differentiating between LS and sporadic CRC in tumors showing loss of MLH1

Reflex pattern – if no BRAF mutation is detected, MLH1 promoter methylation is evaluated

Analytic sensitivity – methylation levels >10% are reported as positive

Limitations 

Mutations other than BRAF V600E will not be detected

Rare false negatives may occur due to primer- and probe-site mutations

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

Recommended Use 

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

This test is also used within the LS reflex testing pathway (for CRC specimens only)

Clinical sensitivity – activating BRAF mutation found in ~10% of CRCs

Analytic sensitivity/ specificity – 100%

Limitations 

Limit of detection

  • MassARRAY and pyrosequencing − 10% mutant alleles
    • MassARRAY and pyrosequencing – oncogenic mutations outside of codon 600 will not be detected
  • NGS – 5% mutant alleles

HNPCC/Lynch Syndrome Deletion/Duplication 2001728
Method: Polymerase Chain Reaction/Multiplex Ligation-dependent Probe Amplification

Recommended Use 

Order if sequencing studies have been performed previously at another laboratory

Order if there is a known familial deletion or duplication

Both sequencing and deletion/duplication testing are necessary to detect all pathogenic mutations in MMR genes

Requires permission from ARUP's Genetic Counselors (800-242-2787, x2141) before ordering

Guidelines

Balmana J, Balaguer F, Cervantes A, Arnold D, ESMO Guidelines Working Group. Familial risk-colorectal cancer: ESMO Clinical Practice Guidelines. Ann Oncol. 2013; 24 Suppl 6: vi73-80. PubMed

Evaluation of Genomic Applications in Practice and Prevention (EGAPP) Working Group. Recommendations from the EGAPP Working Group: genetic testing strategies in newly diagnosed individuals with colorectal cancer aimed at reducing morbidity and mortality from Lynch syndrome in relatives. Genet Med. 2009; 11(1): 35-41. PubMed

Giardiello FM, Allen JI, Axilbund JE, Boland R, Burke CA, Burt RW, Church JM, Dominitz JA, Johnson DA, Kaltenbach T, Levin TR, Lieberman DA, Robertson DJ, Syngal S, Rex DK, US Multi-Society Task Force on Colorectal Cancer. Guidelines on genetic evaluation and management of Lynch syndrome: a consensus statement by the US Multi-Society Task Force on colorectal cancer. Gastroenterology. 2014; 147(2): 502-26. PubMed

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

NCCN Clinical Practice Guidelines in Oncology, Colorectal Cancer Screening. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Nov 2015]

NCCN Clinical Practice Guidelines in Oncology, Genetic/Familial High-Risk Assessment: Colorectal. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Jan 2016]

NCCN Clinical Practice Guidelines in Oncology, Uterine Neoplasms. National Comprehensive Cancer Network. Fort Washington, PA [Accessed: Jul 2015]

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

Rubenstein JH, Enns R, Heidelbaugh J, Barkun A, Clinical Guidelines Committee. American Gastroenterological Association Institute Guideline on the Diagnosis and Management of Lynch Syndrome Gastroenterology. 2015; 149(3): 777-82; quiz e16-7. 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

Vasen HF, Möslein G, Alonso A, Bernstein I, Bertario L, Blanco I, Burn J, Capella G, Engel C, Frayling I, Friedl W, Hes FJ, Hodgson S, Mecklin J, Møller P, Nagengast F, Parc Y, Renkonen-Sinisalo L, Sampson JR, Stormorken A, Wijnen J. Guidelines for the clinical management of Lynch syndrome (hereditary non-polyposis cancer). J Med Genet. 2007; 44(6): 353-62. PubMed

General References

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

Boland R, Goel A. Microsatellite instability in colorectal cancer. Gastroenterology. 2010; 138(6): 2073-2087.e3. PubMed

Djordjevic B, Broaddus RR. Role of the clinical pathology laboratory in the evaluation of endometrial carcinomas for Lynch syndrome. Semin Diagn Pathol. 2014; 31(3): 195-204. PubMed

Geiersbach KB, Samowitz WS. Microsatellite instability and colorectal cancer. Arch Pathol Lab Med. 2011; 135(10): 1269-77. PubMed

Jasperson KW, Tuohy TM, Neklason DW, Burt RW. Hereditary and familial colon cancer. Gastroenterology. 2010; 138(6): 2044-58. PubMed

Legolvan MP, Taliano RJ, Resnick MB. Application of molecular techniques in the diagnosis, prognosis and management of patients with colorectal cancer: a practical approach. Hum Pathol. 2012; 43(8): 1157-68. PubMed

Lynch HT, Lynch PM, Lanspa SJ, Snyder CL, Lynch JF, Boland CR. Review of the Lynch syndrome: history, molecular genetics, screening, differential diagnosis, and medicolegal ramifications. Clin Genet. 2009; 76(1): 1-18. PubMed

Rybak C, Hall MJ. Interpretation of genetic testing for lynch syndrome in patients with putative familial colorectal cancer. J Natl Compr Canc Netw. 2011; 9(11): 1311-20. PubMed

Senter L. Genetic testing by cancer site: colon (nonpolyposis syndromes). Cancer J. 2012; 18(4): 334-7. PubMed

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

References from the ARUP Institute for Clinical and Experimental Pathology®

Chadwick BE. Beyond cytomorphology: expanding the diagnostic potential for biliary cytology. Diagn Cytopathol. 2012; 40(6): 536-41. PubMed

Hegde M, Ferber M, Mao R, Samowitz W, Ganguly A, Working Group of the American College of Medical Genetics and Genomics (ACMG) Laboratory Quality Assurance Committee. ACMG technical standards and guidelines for genetic testing for inherited colorectal cancer (Lynch syndrome, familial adenomatous polyposis, and MYH-associated polyposis). Genet Med. 2014; 16(1): 101-16. 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

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

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

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

Medical Reviewers

Samowitz, Wade S., MD, Medical Director, Solid Tumor Molecular Diagnostics and Histology, and Staff Pathologist, Anatomic Pathology at ARUP Laboratories; Professor of Anatomic Pathology, University of Utah

Samowitz

Last Update: May 2017