Clinical Variant Interpretation

Prioritize and interpret variants for clinical significance using databases and ACMG/AMP guidelines.

Interpretation Framework

Annotated VCF
    │
    ├── Database Lookup
    │   ├── ClinVar (clinical assertions)
    │   ├── OMIM (disease associations)
    │   └── gnomAD (population frequency)
    │
    ├── Computational Predictions
    │   ├── SIFT, PolyPhen-2
    │   ├── CADD, REVEL
    │   └── SpliceAI
    │
    ├── ACMG Classification
    │   └── Pathogenic → Likely Pathogenic → VUS → Likely Benign → Benign
    │
    └── Prioritized Variant List

ClinVar Annotation

Download ClinVar

bash

wget https://ftp.ncbi.nlm.nih.gov/pub/clinvar/vcf_GRCh38/clinvar.vcf.gz
wget https://ftp.ncbi.nlm.nih.gov/pub/clinvar/vcf_GRCh38/clinvar.vcf.gz.tbi

Annotate with bcftools

bash

bcftools annotate \
    -a clinvar.vcf.gz \
    -c INFO/CLNSIG,INFO/CLNDN,INFO/CLNREVSTAT \
    input.vcf.gz -Oz -o with_clinvar.vcf.gz

Filter Pathogenic Variants

bash

# Pathogenic or Likely pathogenic
bcftools view -i 'INFO/CLNSIG~"Pathogenic" || INFO/CLNSIG~"Likely_pathogenic"' \
    with_clinvar.vcf.gz -Oz -o pathogenic.vcf.gz

# Exclude benign
bcftools view -e 'INFO/CLNSIG~"Benign" || INFO/CLNSIG~"Likely_benign"' \
    with_clinvar.vcf.gz -Oz -o not_benign.vcf.gz

ClinVar Significance Levels

CLNSIG	Meaning	Action
Pathogenic	Disease-causing	Report
Likely_pathogenic	Probably disease-causing	Report with caveat
Uncertain_significance	VUS	May report, needs follow-up
Likely_benign	Probably not disease-causing	Usually exclude
Benign	Not disease-causing	Exclude
Conflicting	Multiple interpretations	Manual review

ClinVar Review Status

CLNREVSTAT	Stars	Meaning
practice_guideline	4	Expert panel reviewed
reviewed_by_expert_panel	3	ClinGen expert reviewed
criteria_provided,_multiple_submitters	2	Consistent assertions
criteria_provided,_single_submitter	1	One submitter with criteria
no_assertion_criteria	0	No criteria provided

bash

# Filter for high-confidence assertions (2+ stars)
bcftools view -i 'INFO/CLNREVSTAT~"multiple_submitters" || \
    INFO/CLNREVSTAT~"expert_panel" || \
    INFO/CLNREVSTAT~"practice_guideline"' \
    with_clinvar.vcf.gz -Oz -o high_confidence.vcf.gz

InterVar (ACMG Classification)

Automated ACMG/AMP variant classification.

Installation

bash

git clone https://github.com/WGLab/InterVar.git
cd InterVar
# Download databases per documentation

Run InterVar

bash

python Intervar.py \
    -i input.avinput \
    -o output \
    -b hg38 \
    -d humandb/ \
    --input_type=AVinput

From VCF

bash

# Convert VCF to ANNOVAR format
convert2annovar.pl -format vcf4 input.vcf > input.avinput

# Run InterVar
python Intervar.py -i input.avinput -o intervar_results -b hg38

ACMG/AMP Criteria

Pathogenic Criteria

Code	Type	Description
PVS1	Very Strong	Null variant in gene where LOF is disease mechanism
PS1-4	Strong	Same AA change, functional studies, etc.
PM1-6	Moderate	Hot spot, absent from controls, etc.
PP1-5	Supporting	Co-segregation, computational evidence

Benign Criteria

Code	Type	Description
BA1	Stand-alone	AF >5% in gnomAD
BS1-4	Strong	AF greater than expected, functional studies
BP1-7	Supporting	Missense in gene with truncating mechanism

Population Frequency Filtering

bash

# Rare variants only (gnomAD AF < 0.01)
bcftools view -i 'INFO/gnomAD_AF<0.01 || INFO/gnomAD_AF="."' \
    input.vcf.gz -Oz -o rare.vcf.gz

# Ultra-rare for dominant diseases (AF < 0.0001)
bcftools view -i 'INFO/gnomAD_AF<0.0001 || INFO/gnomAD_AF="."' \
    input.vcf.gz -Oz -o ultrarare.vcf.gz

Pathogenicity Score Filtering

CADD Scores

bash

# CADD > 20 (top 1% deleterious)
bcftools view -i 'INFO/CADD_PHRED>20' input.vcf.gz -Oz -o cadd_filtered.vcf.gz

# CADD > 30 (top 0.1%)
bcftools view -i 'INFO/CADD_PHRED>30' input.vcf.gz -Oz -o highly_deleterious.vcf.gz

REVEL Scores

bash

# REVEL > 0.5 (likely pathogenic)
bcftools view -i 'INFO/REVEL>0.5' input.vcf.gz -Oz -o revel_filtered.vcf.gz

Combined Filtering

bash

bcftools view -i '(INFO/CADD_PHRED>20 || INFO/REVEL>0.5) && \
    (INFO/CLNSIG~"Pathogenic" || INFO/CLNSIG~"Likely" || INFO/CLNSIG=".")' \
    input.vcf.gz -Oz -o prioritized.vcf.gz

Python: Clinical Prioritization

python

from cyvcf2 import VCF, Writer

def classify_variant(variant):
    clnsig = variant.INFO.get('CLNSIG', '')
    af = variant.INFO.get('gnomAD_AF', 0) or 0
    cadd = variant.INFO.get('CADD_PHRED', 0) or 0
    revel = variant.INFO.get('REVEL', 0) or 0

    # Known pathogenic
    if 'Pathogenic' in str(clnsig):
        return 'PATHOGENIC'
    if 'Likely_pathogenic' in str(clnsig):
        return 'LIKELY_PATHOGENIC'

    # Known benign
    if 'Benign' in str(clnsig) or af > 0.05:
        return 'BENIGN'

    # Computational prediction
    if cadd > 25 or revel > 0.7:
        if af < 0.0001:
            return 'LIKELY_PATHOGENIC'
        elif af < 0.01:
            return 'VUS_FAVOR_PATH'

    if cadd < 10 and revel < 0.3:
        return 'LIKELY_BENIGN'

    return 'VUS'

vcf = VCF('annotated.vcf.gz')
results = []

for variant in vcf:
    classification = classify_variant(variant)
    if classification in ('PATHOGENIC', 'LIKELY_PATHOGENIC', 'VUS_FAVOR_PATH'):
        gene = variant.INFO.get('SYMBOL', 'Unknown')
        consequence = variant.INFO.get('Consequence', 'Unknown')
        results.append({
            'chrom': variant.CHROM,
            'pos': variant.POS,
            'ref': variant.REF,
            'alt': variant.ALT[0],
            'gene': gene,
            'consequence': consequence,
            'classification': classification,
            'clnsig': variant.INFO.get('CLNSIG', '.'),
            'cadd': variant.INFO.get('CADD_PHRED', '.'),
            'af': variant.INFO.get('gnomAD_AF', '.')
        })

# Output prioritized variants
for r in results:
    print(f"{r['gene']}\t{r['chrom']}:{r['pos']}\t{r['consequence']}\t{r['classification']}")

Gene Panel Filtering

bash

# Filter to gene panel
bcftools view -R gene_panel.bed input.vcf.gz -Oz -o panel_variants.vcf.gz

# Or by gene symbol (requires VEP annotation)
bcftools view -i 'INFO/CSQ~"BRCA1" || INFO/CSQ~"BRCA2"' \
    input.vcf.gz -Oz -o brca_variants.vcf.gz

Disease-Specific Resources

Resource	Content	Use
ClinVar	Clinical assertions	Primary lookup
OMIM	Gene-disease relationships	Gene prioritization
HGMD	Published mutations	Literature evidence
gnomAD	Population frequencies	Rarity filtering
ClinGen	Gene validity/dosage	LOF interpretation

Reporting Template

bash

bcftools query -f '%CHROM\t%POS\t%REF\t%ALT\t%INFO/SYMBOL\t%INFO/Consequence\t\
%INFO/CLNSIG\t%INFO/CLNDN\t%INFO/gnomAD_AF\t%INFO/CADD_PHRED\n' \
    prioritized.vcf.gz > clinical_report.tsv

Complete Workflow

bash

#!/bin/bash
set -euo pipefail

INPUT=$1
CLINVAR=$2
OUTPUT_PREFIX=$3

echo "=== Add ClinVar annotations ==="
bcftools annotate -a $CLINVAR \
    -c INFO/CLNSIG,INFO/CLNDN,INFO/CLNREVSTAT,INFO/CLNVC \
    $INPUT -Oz -o ${OUTPUT_PREFIX}_clinvar.vcf.gz

echo "=== Filter rare variants ==="
bcftools view -i 'INFO/gnomAD_AF<0.01 || INFO/gnomAD_AF="."' \
    ${OUTPUT_PREFIX}_clinvar.vcf.gz -Oz -o ${OUTPUT_PREFIX}_rare.vcf.gz

echo "=== Extract pathogenic/likely pathogenic ==="
bcftools view -i 'INFO/CLNSIG~"athogenic"' \
    ${OUTPUT_PREFIX}_rare.vcf.gz -Oz -o ${OUTPUT_PREFIX}_pathogenic.vcf.gz

echo "=== Extract high-impact VUS ==="
bcftools view -i 'INFO/CLNSIG~"Uncertain" && INFO/CADD_PHRED>20' \
    ${OUTPUT_PREFIX}_rare.vcf.gz -Oz -o ${OUTPUT_PREFIX}_vus_review.vcf.gz

echo "=== Generate report ==="
bcftools query -H -f '%CHROM\t%POS\t%REF\t%ALT\t%INFO/SYMBOL\t%INFO/Consequence\t\
%INFO/CLNSIG\t%INFO/CLNDN\t%INFO/gnomAD_AF\t%INFO/CADD_PHRED\n' \
    ${OUTPUT_PREFIX}_pathogenic.vcf.gz > ${OUTPUT_PREFIX}_report.tsv

echo "=== Complete ==="
echo "Pathogenic: ${OUTPUT_PREFIX}_pathogenic.vcf.gz"
echo "VUS for review: ${OUTPUT_PREFIX}_vus_review.vcf.gz"
echo "Report: ${OUTPUT_PREFIX}_report.tsv"

Related Skills

variant-calling/variant-annotation - VEP/SnpEff annotation
variant-calling/filtering-best-practices - Quality filtering
database-access/entrez-fetch - Download ClinVar/OMIM data
pathway-analysis/go-enrichment - Gene set analysis

Search AI Tools

bio-variant-calling-clinical-interpretation

Install this agent skill to your Project

SKILL.md

Clinical Variant Interpretation

Interpretation Framework

ClinVar Annotation

Download ClinVar

Annotate with bcftools

Filter Pathogenic Variants

ClinVar Significance Levels

ClinVar Review Status

InterVar (ACMG Classification)

Installation

Run InterVar

From VCF

ACMG/AMP Criteria

Pathogenic Criteria

Benign Criteria

Population Frequency Filtering

Pathogenicity Score Filtering

CADD Scores

REVEL Scores

Combined Filtering

Python: Clinical Prioritization

Gene Panel Filtering

Disease-Specific Resources

Reporting Template

Complete Workflow

Related Skills