What lab tests are used to diagnose familial hypercholesterolemia?

Laboratory Testing for Familial Hypercholesterolemia

The cornerstone laboratory test for diagnosing familial hypercholesterolemia is a fasting lipid profile measuring LDL-cholesterol, with genetic testing serving as the most accurate confirmatory method when available. 1, 2

Primary Phenotypic Testing

Lipid Profile Components

• LDL-cholesterol measurement is the essential screening test, using age-specific, sex-specific, and country-specific thresholds above the 95th percentile to identify index cases 2
• The complete fasting lipid profile should include total cholesterol, LDL-cholesterol, HDL-cholesterol, and triglycerides 2
• Non-fasting samples may be acceptable for screening purposes, though fasting samples remain preferred 2

Special Considerations for LDL-Cholesterol Measurement

• When triglycerides exceed 4.5 mmol/L (400 mg/dL), use direct LDL-cholesterol assay rather than calculated values, as the Friedewald equation becomes unreliable 1, 2
• For triglycerides between 4.5-10.0 mmol/L (400-850 mg/dl), either measure LDL-cholesterol directly after therapeutic lowering of triglycerides to <4.5 mmol/L, or use novel calculation equations 1
• Adjust LDL-cholesterol values for concurrent lipid-lowering medications (statins, ezetimibe, PCSK9 inhibitors) when making a phenotypic diagnosis, particularly if reliable pretreatment values are unavailable 1, 2
• Repeat testing after full recovery from acute illness if the diagnosis remains uncertain, as acute conditions can temporarily alter lipid levels 1, 2

Genetic Testing (Gold Standard)

Primary Gene Panel

Genetic testing should use targeted next-generation sequencing of all exons and exon-intron boundaries of four key genes: 1

• LDLR (LDL receptor gene) - most common cause
• APOB (apolipoprotein B gene) - specifically the exons encoding the LDLR ligand-binding region
• PCSK9 (proprotein convertase subtilisin/kexin type 9 gene)
• LDLRAP1 (LDL receptor adaptor protein 1 gene)

Technical Requirements

• Testing must include analysis for deletions and duplications in LDLR, not just point mutations 1
• Genetic testing should be performed in an accredited, certified laboratory using standardized methods 1
• Variants must be classified according to ACMG, AMP, or ClinGen FH Variant Curation Expert Panel guidelines 1

When to Order Genetic Testing

• Offer genetic testing to all individuals with strong clinical suspicion based on phenotypic criteria (definite or highly probable heterozygous FH, or phenotypic homozygous FH) 1
• Consider genetic testing in those with probable phenotypic diagnosis of heterozygous FH 1
• May consider in possible FH cases when incomplete information exists and genetic results would affect clinical management 1

Supplementary Tests

Apolipoprotein B (ApoB)

• ApoB measurement may be useful in patients with hypertriglyceridemia, as it estimates the number of atherogenic lipoprotein particles 2
• Remains accurate in non-fasting states but carries additional expense and may have variable laboratory reliability 2

Lipoprotein(a) [Lp(a)]

• Consider testing in those with family history of premature ASCVD or personal history of ASCVD not explained by major risk factors 2
• Lp(a) levels can influence diagnostic accuracy and risk stratification 1

Age-Specific Testing Protocols

Children and Adolescents

• For suspected homozygous FH: test as early as possible (at newborn stage or by age 2 years) 2
• For at-risk heterozygous FH: screen at or after age 5 using LDL-cholesterol, or as early as age 2 in those with strong family history of premature ASCVD 2
• Universal screening using age-specific and sex-specific LDL-cholesterol criteria should be considered for all children and adolescents 2

Adults

• Selective screening should target adults with premature ASCVD and family history of premature ASCVD and/or hypercholesterolemia 2
• Opportunistic screening using LDL-cholesterol >4.9 mmol/L (≥190 mg/dL) should identify community cases 2

Cascade Testing Strategy

After Index Case Identification

Once a pathogenic variant is identified in the proband, cascade genetic testing for that specific variant should be offered sequentially: 1

1. All first-degree relatives first
2. If first-degree relatives are unavailable or decline, proceed to second-degree relatives
3. Then third-degree relatives
4. Continue until all at-risk family members have been offered testing

When Genetic Testing Unavailable

• Use phenotypic cascade testing with age-specific, sex-specific, and country-specific LDL-cholesterol concentrations 1
• Do not use clinical diagnostic tools designed for probands (Dutch Lipid Clinic Network criteria, Simon Broome criteria) for cascade testing of relatives, as these are not validated for this purpose 1

Critical Pitfalls to Avoid

Diagnostic Errors

• Never exclude FH based solely on negative genetic testing if the clinical phenotype strongly suggests FH, as undetected genetic variants may be responsible 1
• Polygenic scores for hypercholesterolemia are not yet fully standardized and should be used with caution 1, 3
• Clinical diagnostic criteria (Dutch Lipid Clinic Network, Simon Broome) become less accurate in patients already on statin therapy 1

Laboratory Interpretation Errors

• The Friedewald equation for calculating LDL-cholesterol becomes unreliable with elevated triglycerides and should not be used when triglycerides exceed 4.5 mmol/L (400 mg/dL) 1, 2
• Failure to adjust for lipid-lowering medications can lead to underdiagnosis 1, 2

Genetic Testing Limitations

• Genetic testing is currently underutilized due to costs, inadequate clinical skills in genomic medicine, genetic privacy concerns, and limited availability of genetic counseling services 3
• Pre-test and post-test genetic counseling should be offered to all patients and at-risk relatives as an integral component of testing 1, 3