What is the recommended evaluation and management when immunohistochemistry shows loss of MLH1, MSH2, and MSH6 expression?

Loss of MLH1, MSH2, and MSH6 Expression: Evaluation and Management

When immunohistochemistry shows loss of MLH1, MSH2, and MSH6 expression simultaneously, this represents an unusual pattern requiring systematic evaluation for Lynch syndrome, starting with MLH1 promoter methylation testing and BRAF V600E mutation analysis to exclude sporadic cancer, followed by germline genetic testing if these are negative or absent. 1, 2

Initial Triage Based on MLH1 Loss

Since MLH1 loss is present, the first critical step is distinguishing sporadic from hereditary causes:

• Test for MLH1 promoter hypermethylation OR BRAF V600E mutation 1, 2, 3
  • If MLH1 promoter is hypermethylated OR BRAF V600E mutation is present → likely sporadic cancer, no germline testing needed
  • If both are negative or not performed → proceed to germline genetic testing 1

Important caveat: BRAF mutation testing is NOT reliable in endometrial or other gynecologic cancers (unlike colorectal cancer) and should not be used as a surrogate for MLH1 methylation in these tissues 4

Understanding This Unusual Pattern

The simultaneous loss of MLH1, MSH2, and MSH6 is extremely rare (approximately 1.4% of all cases, 5.1% of dMMR cases) 5. This pattern suggests:

• Widespread dysregulation of DNA repair processes beyond typical Lynch syndrome
• Possible multiple germline variants affecting different MMR genes simultaneously 5, 6
• Potential secondary mutations in homologous recombination repair genes (ATM, BRCA1, BRCA2, PALB2, etc.) 5

Normal MMR proteins function as heterodimers (MLH1/PMS2 and MSH2/MSH6). Loss of three proteins across both heterodimers indicates either:

1. Multiple pathogenic variants
2. Downstream effects from one primary defect affecting protein stability of others
3. Somatic mutations accumulating due to severe MMR deficiency 5

Germline Genetic Testing Strategy

If MLH1 methylation/BRAF testing excludes sporadic cancer, proceed with comprehensive germline testing 1, 7, 2:

1. Begin testing with the gene corresponding to the absent protein based on IHC results 7

   • In this case: Test MLH1 first (since MLH1 is lost)
   • Then test MSH2 and MSH6 sequentially

2. Full germline testing must include 1:

   • DNA sequencing
   • Large rearrangement analysis
   • Consider EPCAM deletion testing (can cause MSH2 loss through hypermethylation) 1, 8

3. Consider expanded testing for homologous recombination repair genes given the unusual triple-loss pattern 5

Genetic Counseling Referral

All patients with loss of MSH2, MSH6, or PMS2 should be referred for genetic counseling 9, 10. For MLH1 loss, refer if:

• Age <60 years at diagnosis 1, 9
• Multiple Lynch syndrome-associated cancers
• First-degree relative with colorectal or endometrial cancer 9

Critical barrier to address: Only 38% of eligible patients are actually referred for genetic counseling, and only 20.6% complete germline testing 11. The primary barrier is lack of provider referral 11. Consider:

• Automated referral systems
• Navigation support
• Direct communication with genetics team

Surveillance if Lynch Syndrome Confirmed

Colonoscopy surveillance 1, 7, 8:

• Begin at age 20-25 years (or 10 years before youngest family diagnosis)
• Repeat every 1-2 years (evidence shows 62-72% reduction in CRC mortality) 1, 8
• More frequent surveillance (every 1-2 years vs. every 3 years) associated with earlier stage detection 8

Gynecologic surveillance 1, 7:

• Annual examination starting age 30-35 years
• Transvaginal ultrasound, CA-125, endometrial biopsy 1
• Prophylactic hysterectomy/bilateral salpingo-oophorectomy after childbearing is complete (lifetime endometrial cancer risk up to 60%) 9, 7, 10

Upper GI surveillance 7:

• Test for and eradicate H. pylori 1
• Consider upper endoscopy every 1-3 years if high gastric cancer incidence in family 1, 7

Chemoprevention 1:

• Aspirin 600 mg daily for at least 2 years shows 60% reduction in Lynch syndrome-associated cancers 1

Genotype-Phenotype Correlations

Understanding which gene is mutated helps predict cancer risk 1:

• MLH1 mutations: Higher risk of young-onset CRC
• MSH2 mutations: Higher risk of extracolonic cancers
• MSH6 mutations: Increased endometrial cancer risk, later age of onset (consider starting colonoscopy at 30-35 years) 7
• PMS2 mutations: Lower absolute lifetime CRC/endometrial cancer risk (15-20%) 1

Family Testing

Once a germline mutation is identified:

• Test first-degree relatives for the specific familial mutation 7
• If first-degree relatives unavailable/unwilling, offer testing to more distant relatives 7
• Mutation-negative relatives can follow average-risk screening 7