Variant classification summary

NM_000143.4:c.817G>A

FH · NP_000134.2:p.(Ala273Thr) · NM_000143.4

GRCh37: chr1:241669390 C>T · GRCh38: chr1:241506090 C>T

Gene: FH Transcript: NM_000143.4

Final call

Likely Pathogenic

PS3 moderate PM1 moderate PM2 moderate PP1 supporting PP2 supporting PP3 supporting PP4 supporting PP5 supporting

Variant details

Gene

FH

Transcript

NM_000143.4

Protein

NP_000134.2:p.(Ala273Thr)

gnomAD AF

3.098200565111783e-06 (v4.1)

ClinVar

Pathogenic

OncoKB

Unknown Oncogenic Effect

Classification rationale

Interpretation summary

Generated evidence synthesis

1

FH c.817G>A (p.Ala273Thr) is a missense variant in exon 6 of the fumarate hydratase gene, located within the fumarate lyase catalytic domain.

2

The variant is extremely rare in population databases: gnomAD v2.1 allele frequency 0.00040% (1/251,224 alleles) and v4.1 allele frequency 0.00031% (5/1,613,840 alleles), with no homozygotes observed (PM2).

3

Functional studies from patient tumor tissue demonstrate loss of fumarate hydratase function: immunohistochemistry showed diffuse/strong positive 2SC staining, negative FH protein staining, and decreased 5-hmC, with loss of heterozygosity confirmed in tumor tissue (PS3_Moderate).

4

The variant resides in the fumarate lyase domain, the critical catalytic domain of FH, and has been observed in 2 of 13 FH-mutated PPGL cases across the literature suggesting recurrence at this site (PM1).

5

Co-segregation was observed in two affected family members: the proband with bladder paraganglioma and the proband's aunt who carried the same variant and developed bladder paraganglioma in her 20s (PP1).

6

Multiple lines of in silico evidence support a deleterious effect: REVEL score 0.867 strongly predicts damaging impact (PP3).

7

The patient's phenotype of bladder paraganglioma with a positive family history is highly specific for hereditary PPGL syndromes including FH-related disease (PP4).

8

FH is a well-established disease gene where missense variants are a known pathogenic mechanism in HLRCC and PPGL (PP2).

9

This variant has been reported in ClinVar as Pathogenic (Variation ID: 214377), supported by multiple independent clinical laboratory submissions (PP5).

Final determination: Generic ACMG/AMP 2015 fallback rules support a Likely Pathogenic classification based on the observed combination of pathogenic criteria.

Criteria assessment

ACMG/AMP criteria review

Criteria shown when status is available

All criteria require review: For research and educational purposes only.

Criterion	Status	Rationale	Evidence used
PVS1	N/A	This is a missense substitution (c.817G>A, p.Ala273Thr) in exon 6. The variant does not fall into the default generic PVS1 null-variant buckets of nonsense, frameshift, or canonical ±1,2 splice consensus variants. Generic PVS1 framework is not applicable to missense variants.	pvs1_gene_context pvs1_variant_assessment
PS1	N/A	No prior pathogenic variant at the same amino acid position with a different nucleotide change has been established in the case materials.
PS2	Not met	No de novo evidence with confirmed maternity and paternity was identified for this variant in the available literature or ClinVar submissions.
PS3	Met	Variant-specific functional evidence from one publication (Ma et al. 2022, PMID:35821608) demonstrates loss of fumarate hydratase function: immunohistochemistry of the patient's tumor tissue showed diffuse/strong positive staining for 2-succinocysteine (2SC) and negative staining for FH protein, with decreased 5-hydroxymethylcytosine (5-hmC), confirming loss of enzymatic activity and a hypermethylation phenotype. Loss of heterozygosity was also demonstrated in tumor tissue. A single study with direct variant-specific functional characterization supports moderate-strength PS3.	PMID:35821608
PS4	Not met	The variant was observed in 1 of 319 PPGL patients (PMID:35821608), which does not provide statistically significant enrichment over the extremely low population frequency. Multiple ClinVar submissions from clinical laboratories suggest clinical observation but do not constitute controlled case-control data suitable for PS4.	PMID:35821608 clinvar gnomad_v2 gnomad_v4
PS5	N/A	PS5 is not a standard criterion in the ACMG/AMP 2015 framework for germline variant interpretation.
PM1	Met	The variant is located at codon 273 within the fumarate lyase domain of FH, the catalytic domain responsible for enzymatic conversion of fumarate to malate. This is a well-characterized critical functional domain. The variant causes a missense change (Ala273Thr) within this domain, and functional data confirm loss of enzyme activity. Although the residue is not a statistically significant hotspot in cancerhotspots.org, the well-established functional domain supports PM1 at moderate strength.	PMID:35821608
PM2	Met	The variant is extremely rare in population databases: gnomAD v2.1 allele frequency = 0.00040% (1/251,224 alleles, 0 homozygotes), gnomAD v4.1 allele frequency = 0.00031% (5/1,613,840 alleles, 0 homozygotes), and absent from gnomAD-Canada. The highest subpopulation frequency is 0.0032% (gnomAD v4.1, Remaining individuals), which is well below the 0.1% PM2 threshold for non-VCEP application. The variant is absent or extremely rare in all populations surveyed.	gnomad_v2 gnomad_v4 gnomad_canada
PM5	Not met	No pathogenic missense variant at the same codon (Ala273) with a different amino acid change was identified in the PM5 candidate search. The automated pm5_candidates pipeline found no same-residue comparator variants.	pm5_candidates
PM6	Not met	No evidence of assumed or confirmed de novo occurrence was identified for this variant. The variant was observed with a positive family history (aunt with same variant and bladder PGL in PMID:35821608), arguing against a de novo event.	PMID:35821608
PP1	Met	Co-segregation observed in one affected family member: the proband's aunt carried the same c.817G>A variant and developed bladder paraganglioma in her 20s (PMID:35821608). This represents co-segregation in two affected relatives, supporting PP1 at supporting strength.	PMID:35821608
PP2	Met	FH is a tumor suppressor gene where missense variants are a well-established mechanism of disease. Both truncating and missense pathogenic variants in FH cause hereditary leiomyomatosis and renal cell cancer (HLRCC) as well as pheochromocytoma/paraganglioma. The gene has a low rate of benign missense variation relative to pathogenic missense changes, as evidenced by the extreme rarity of this variant in gnomAD and the known disease association of FH missense variants.	PMID:35821608 gnomad_v2 gnomad_v4
PP3	Met	Multiple lines of in silico evidence support a deleterious effect: REVEL score of 0.867 (strongly predicted damaging), and BayesDel score of 0.503 (borderline). SpliceAI predicts no splicing impact (max delta = 0.00). The high REVEL score is the primary computational support. The missense change from alanine to threonine at a position within the catalytic fumarate lyase domain is predicted to disrupt protein function.	revel bayesdel spliceai
PP4	Met	The patient presented with bladder paraganglioma, a phenotype highly specific for hereditary PPGL syndromes including FH-related disease. The presence of a positive family history of PPGL (affected aunt) further supports a hereditary etiology. The clinical phenotype is consistent with FH-associated disease.	PMID:35821608
PP5	Met	The variant is classified as Pathogenic in ClinVar (Variation ID: 214377), supported by multiple independent clinical laboratory submissions (3 labs report Pathogenic, 2 labs report Likely pathogenic/likely pathogenic, 1 lab reports Uncertain significance). Multiple reputable clinical testing laboratories have independently classified this variant as pathogenic or likely pathogenic. Although the ClinVar aggregate review status is 'criteria provided, single submitter' (not 3-star expert panel), the multiple independent clinical classifications constitute reputable source evidence for PP5 at supporting strength under generic ACMG/AMP 2015.	clinvar
BA1	Not met	The maximum population allele frequency (gnomAD v4.1, Remaining individuals: 0.0032%) is far below the 1% BA1 threshold for non-VCEP application.	gnomad_v2 gnomad_v4
BS1	Not met	The maximum population allele frequency (gnomAD v4.1: 0.0032%) is far below the 0.3% BS1 threshold for non-VCEP application.	gnomad_v2 gnomad_v4
BS2	Not met	No evidence of this variant being observed in a healthy adult individual with full penetrance expected at an early age. The variant is ultra-rare in gnomAD and has only been observed in affected individuals.
BS3	Not met	Functional evidence from PMID:35821608 demonstrates a damaging effect: IHC shows loss of FH protein expression with positive 2SC staining and decreased 5-hmC, confirming loss of fumarate hydratase function. This directly contradicts BS3.	PMID:35821608
BS4	Not met	No evidence of lack of segregation in affected family members. The available segregation data (PMID:35821608) shows co-segregation in two affected relatives, which supports pathogenicity rather than BS4.	PMID:35821608
BP1	Not met	FH is a gene where both missense and truncating variants are established causes of disease (HLRCC and PPGL). Missense variants in FH are known to be pathogenic, as demonstrated by multiple pathogenic FH missense variants reported in the literature (PMID:35821608 Table 2). BP1 does not apply.	PMID:35821608
BP2	Not met	No evidence of this variant being observed in trans with a known pathogenic FH variant. FH-related disease is autosomal dominant, and biallelic FH variants cause fumarase deficiency, a distinct severe recessive disorder. No such data available.
BP4	Not met	Computational evidence does not suggest a benign impact. REVEL score of 0.867 strongly predicts a damaging effect. SpliceAI shows no splicing impact (max delta = 0.00), but this does not outweigh the strong deleterious prediction from REVEL. BP4 requires multiple lines of computational evidence suggesting no impact, which is not satisfied.	revel bayesdel spliceai
BP5	Not met	No evidence that this variant was found in a case with an alternate molecular basis for disease. Available data show the variant in individuals with phenotypes consistent with FH-related disease (PPGL, family history of PPGL).
BP6	Not met	ClinVar overall classification for this variant is Pathogenic (Variation ID: 214377). No reputable source has classified this variant as benign or likely benign. BP6 does not apply.	clinvar
BP7	N/A	This is a missense variant (c.817G>A, p.Ala273Thr), not a synonymous variant. BP7 applies only to synonymous variants without predicted splicing impact.

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