ClinVar Genomic variation as it relates to human health

Variant summary: BTD c.1270G>C (p.Asp424His), also known as c.1330G>C (p.Asp444His), results in a non-conservative amino acid change located in the Domain B (biotin-binding, Borsatto_2019) of the encoded protein sequence. Five of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 0.032 in 251430 control chromosomes in the gnomAD database, including 183 homozygotes. The observed variant frequency is significantly higher than estimated for a pathogenic variant in BTD causing Biotinidase Deficiency phenotype, suggesting that the variant is benign. c.1270G>C has been reported in the literature in multiple individuals affected with partial Biotinidase deficiency, a milder form of Biotinidase deficiency that is amenable to treatment with biotin (example, Swango_1998, Wiltink_2016, Procter_2016, Yilmaz_2018). Many of the patients presenting with partial Biotinidase deficiency attributed to this variant (c.1270G>C) were compound heterozygous with other BTD variants associated with a mild or a severe phenotype. However, this variant has also been observed in reportedly normal homozygous individuals who had >30% of wild-type levels of serum Biotinidase enzyme activity (Wiltink_2016). Since serum Biotinidase activity levels >30% are considered normal (Wiltnik_2016), these findings imply that homozygotes for this variant will be unaffected, a finding that is consistent with the high frequency of homozygotes in gnomAD database. c.1270G>C has also been observed in cis with p.Ala151Thr (also known as p.Ala171Thr) forming a complex allele that causes less than 10% of mean normal activity or profound Biotinidase deficiency in that allele (PMID: 10206677, Swango_1998, Wiltink_2016). Multiple publications report experimental evidence evaluating an impact on protein function. One study reported that this variant results in decreased protein expression but does not alter the activity of BTD enzyme (Liu_2018). Another study reports 46% of wild-type activity in cell homogenates but not in culture medium suggestive of an effect on enzyme lability rather than enzyme activity (Borsatto_2019). It is unclear how a 46% activity in cell homogenates correlates with a >30% activity in serum, which is considered normal. The authors state that the mechanism of decreased activity for this variant remains speculative (Borsatto_2019). Twenty ClinVar submitters (evaluation after 2014) cite the variant as uncertain significance (n=2), likely pathogenic (n=1) and pathogenic (n=17). Based on the evidence outlined above, the variant was classified as pathogenic in association with the phenotype of partial Biotinidase deficiency, a condition that is actionable and amenable to treatment with biotin.

ACMG categories: PS3,PM3,PM7,PP3,PP5,BP1

The p.(Asp444His) variant is a well-established hypomorphic pathogenic variant (PMID: 20301497), reported in multiple individuals and classified as pathogenic by multiple laboratories (ClinVar). It is associated with partial biotinidase deficiency when seen in trans with a pathogenic variant.

his variant has been reported in the published literature and is associated with partial biotinidase deficiency (10-30% of normal BTD activity) when a BTD pathogenic variant associated with profound biotinidase deficiency is present on the opposite chromosome (i.e., variants are present in trans) (PMID: 9654207 (1998), 10206677 (1998), 12227467 (2002), 23644139 (2013), 28498829 (2017)). Individuals who are homozygous for this variant are usually asymptomatic and have approximately 50% of normal BTD enzyme activity (PMID: 20539236 (2010), 20556795 (2010), 28682309 (2017)). Based on the available information, this variant is classified as pathogenic

The p.Asp424His variant (also reported as p.Asp444His in the literature) in the BTD gene is a well reported cause of partial biotinidase deficiency (Canda et al., 2018; Swango et al., 1998). This variant in the homozygous state does not cause biotinidase deficiency; however, this variant in conjunction with another profound disease-causing variant is a common cause of partial biotinidase deficiency. This variant has been reported in individuals affected with profound biotinidase deficiency when this variant is in cis with the p.Ala171Thr variant and in trans with a third variant. The p.Asp424His variant was determined to be in trans with multiple pathogenic variants (including p.Cys33Phefs*36), consistent with autosomal recessive inheritance. The presence of this variant with a disease-causing variant on the opposite allele increases suspicion for its pathogenicity. The p.Asp424His variant has also been identified in 5,119/129,150 European chromosomes by the Genome Aggregation Database (http://gnomad.broadinstitute.org/). Although this variant has been seen in the general population, its frequency is consistent with a recessive carrier frequency for a mild allele. Well-established in vitro functional studies and patient assays of the p.Asp424His variant demonstrate reduced enzymatic activity that is sufficient to be disease-causing (Liu et al., 2018; Swango et al., 1998). Computational tools predict that this variant is deleterious; however, the accuracy of in silico algorithms is limited. These data were assessed using the ACMG/AMP variant interpretation guidelines. In summary, there is sufficient evidence to classify the p.Asp424His variant as pathogenic for autosomal recessive biotinidase deficiency based on the information above. [ACMG evidence codes used: PS3; PM3_strong; PP3; PP4]

The BTD c.1270G>C; p.Asp424His variant (rs13078881), also known as c.1330G>C; p.Asp444His for NM_000060.2, is reported in multiple patients with partial biotinidase deficiency (Dobrowolski 2003, Funghini 2002, Milankovics 2010, Muhl 2001, Pomponio 2000, Swango 1998, Wolf 2005), with a higher prevalence in affected individuals (Milankovics 2010). This variant is reported in ClinVar (Variation ID: 1900), and found in the general population with an overall allele frequency of 3.2% (9005/282830 alleles, including 199 homozygotes) in the Genome Aggregation Database. The aspartate at codon 424 is highly conserved, and computational analyses predict that the variant is deleterious (REVEL: 0.769). Based on available information, this variant is considered to be mildly pathogenic. References: Dobrowolski S et al. Real time PCR assays to detect common mutations in the biotinidase gene and application of mutational analysis to newborn screening for biotinidase deficiency. Mol Genet Metab. 2003 Feb;78(2):100-7. PMID: 12618081 Funghini S et al. Two new mutations in children affected by partial biotinidase deficiency ascertained by newborn screening. J Inherit Metab Dis. 2002 Aug;25(4):328-30. PMID: 12227467. Milankovics I et al. High frequencies of biotinidase (BTD) gene mutations in the Hungarian population. J Inherit Metab Dis. 2010 Dec;33 Suppl 3:S289-92. PMID: 20549359. Muhl A et al. Molecular characterisation of 34 patients with biotinidase deficiency ascertained by newborn screening and family investigation. Eur J Hum Genet. 2001 Apr;9(4):237-43. PMID: 11313766. Pomponio R et al. Novel mutations cause biotinidase deficiency in Turkish children. J Inherit Metab Dis. 2000 Mar;23(2):120-8. PMID: 10801053. Swango K et al. Partial biotinidase deficiency is usually due to the D444H mutation in the biotinidase gene. Hum Genet. 1998 May;102(5):571-5. PMID: 9654207. Wolf B et al. Biotinidase deficiency: novel mutations and their biochemical and clinical correlates. Hum Mutat. 2005 Apr;25(4):413. PMID: 9375914.

The p.Asp424His variant (also referred to as p.Asp444His) in BTD has been reported in several individuals with biotinidase deficiency, segregated with disease in affected relatives, and is predicted to lead to a ~25% reduction in biotinidase activity (Norrgard 1998 PMID: 9232193). This variant has also been reported by other clinical laboratories in ClinVar (Variation ID 1900) and has been identified in 5.53% (587/10610) of Finnish chromosomes, including 25 homozygotes, by gnomAD (http://gnomad.broadinstitute.org, v.3.1.2). Although this variant is seen in the general population, its frequency is consistent with its predicted effect. Individuals homozygous for the p.Asp424His variant are expected to have approximately 50% of mean normal serum biotinidase deficiency, similar to the activity of heterozygotes for profound biotinidase deficiency. When found in the compound heterozygous state with a severe pathogenic variant in BTD, this variant is reported to be the most common cause of partial biotinidase deficiency (Swango 1998 PMID: 9654207). In vitro functional studies support an impact on protein function (Borsatto 2019 PMID: 31337602, Liu 2018 PMID: 29359854). In summary, this variant meets criteria to be classified as pathogenic for autosomal recessive biotinidase deficiency, though it is considered a mild allele. ACMG/AMP Criteria applied: PM3_Strong, PP1_Strong, PS3_Supporting.

The c.1330G>C (p.D444H) alteration is located in exon 4 (coding exon 4) of the BTD gene. This alteration results from a G to C substitution at nucleotide position 1330, causing the aspartic acid (D) at amino acid position 444 to be replaced by a histidine (H). Based on data from the Genome Aggregation Database (gnomAD), the BTD c.1330G>C alteration was observed in 3.18% (9005/282830) of total alleles studied, with a frequency of 5.56% (1396/25118) in the European (Finnish) subpopulation. This alteration has been reported in essentially all patients with partial biotinidase deficiency (defined as serum biotinidase activity of 10-30%) who have a second profound allele in trans configuration. The D444H allele results in approximately 50% reduction of biotinidase enzyme activity. Heterozygous carriers of D444H have approximately 75% mean normal enzyme activity. Homozygotes for D444H alone have approximately 50% of mean normal enzyme activity, similar to heterozygous carriers of one profound allele, and do not have clinical biotinidase deficiency. However, the double mutation of D444H in cis with A171T results in a profound deficiency allele with <10% of residual enzyme activity. This double mutation has been reported in patients with profound biotinidase deficiency when in trans with another profound allele or in the homozygous state (Swango, 1998; Norrgard, 1998; Mil&aacute;nkovics, 2010; Cowan, 2010). This amino acid position is not well conserved in available vertebrate species. In vitro studies showed slightly reduced but not complete loss of biotinidase activity in HEK293 cells (Borsatto, 2019). This alteration is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic.

The BTD c.1330G>C (p.Asp444His) variant has been reported as the most common variant found in newborns screened for profound biotinidase deficiency. However, in individuals with profound biotinidase deficiency, the variant is usually found in cis with one of three additional variants (Norrgard et al. 1999). In the studies by Norrgard et al. (1998; 1999), 14 of 31 children with biotinidase deficiency carried both the p.Asp444His and the p.Ala171Thr variants in cis. This double variant allele has been identified as the second most common allele in newborns screened for biotinidase deficiency. The combination of the two variants resulted in approximately 52% enzyme loss. In control populations, 23 of 296 healthy individuals were identified with the p.Asp444His variant versus none of 376 with the p.Ala171Thr variant. The p.Asp444His variant alone is reported to be the most common cause of partial biotinidase deficiency (10% to 30% of normal serum activity) when found in a compound heterozygous state with a severe pathogenic variant in the BTD gene (Swango et al. 1998). Individuals who are homozygous for the p.Asp444His variant have approximately 50% of mean normal enzyme activity (Pindolia et al. 2010). Untreated individuals with partial biotinidase deficiency are often asymptomatic in the absence of confounding factors such as significant illness. The p.Asp444His variant is reported at a frequency of 0.0758 in the Finnish population of the 1000 Genomes Project. Although this allele frequency appears inconsistent with the disease prevalence, the p.Asp444His variant appears to be a mild variant when found in trans with a severe pathogenic variant, and is only associated with severe biotinidase deficiency when found in cis with a more severe variant and in trans with a severe pathogenic variant. Based on the evidence, the p.Asp444His variant is classified as pathogenic for biotinidase deficiency. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.

The c.1270G>C;p.(Asp424His) missense variant has been observed in affected individual(s) and ClinVar contains an entry for this variant (ClinVar ID: 1900; PMID: 29353266; 29359854; 27845546; 9654207;10206677;11668630) - PS4. Well-established in vitro or in vivo functional studies support a damaging effect on the gene or gene product (PMID: 31208052; 31337602) - PS3_moderate. The p.(Asp424His) was detected in trans with a pathogenic variant (PMID: 11313766; 9654207; 10206677) - PM3. Multiple lines of computational evidence support a deleterious effect on the gene or gene product - PP3. Patient’s phenotype is highly specific for a disease with a single genetic etiology - PP4 and allele frequency is greater than expected for disorder -BS1. The variant was observed in trans with a pathogenic variant for a fully penetrant dominant gene/disorder or observed in cis with a pathogenic variant in any inheritance pattern (cis with rs13073139) - BP2. In summary, the currently available evidence indicates that the variant is pathogenic.

This is a nonsynonymous variant in the BTD gene (OMIM 609019). Biallelic pathogenic variants in this gene have been associated with autosomal recessive biotinidase deficiency (BTD deficiency). This missense variant, also known as p.Asp444His, is a common pathogenic variant associated with partial biotinidase deficiency when found in trans with a more severe pathogenic variant (PMID: 20301497) (PM3_Very Strong). Functional studies have shown that this variant alters BTD protein function (PMID: 29359854, 31337602) (PS3_Moderate). Multiple computational algorithms predict a deleterious effect for this substitution (PP3). This variant has a 4.031% maximum allele frequency in non-founder control populations (https://gnomad.broadinstitute.org/). Based on the current evidence, this variant is classified as pathogenic for autosomal recessive BTD deficiency.

Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Pathogenic. Following criteria are met: 0102 - Loss of function is a known mechanism of disease in this gene and is associated with biotinidase deficiency (MIM#253260). (I) 0106 - This gene is associated with autosomal recessive disease. (I) 0115 - Variants in this gene are known to have variable expressivity. The condition associated with this gene is known to range from partial, with milder symptoms and later onset, to profound, with more severe symptoms and earlier onset. Also, individuals with the same genotype have been observed to have different clinical and biochemical phenotypes (PMIDs: 20301497, 28498829). (I) 0200 - Variant is predicted to result in a missense amino acid change from aspartic acid to histidine. (I) 0252 - This variant is homozygous. (I) 0307 - Variant is present in gnomAD (v2) at a frequency >=0.05 (8607 heterozygotes, 199 homozygotes). (SB) 0309 - An alternative amino acid change at the same position has been observed in gnomAD (v2) (1 heterozygote, 0 homozygotes). (I) 0502 - Missense variant with damaging in silico predictions and low conservation. (I) 0600 - Variant is located in the annotated vanin C-terminal domain (NCBI). (I) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This variant has been observed in multiple individuals with biotinidase deficiency and is described as a mild variant. This variant is not expected to cause disease when homozygous, but is known to cause mild or partial biotinidase dificiency when in trans with a severe pathogenic variant, and can cause profound deficiency when also in cis with a modifying variant (typically p.(Ala151Thr)) (ClinVar, PMIDs: 31337602, 28498829, 28682309). (SP) 1002 - This variant has moderate functional evidence supporting abnormal protein function. Western blot and enzyme activity assays for this variant have been variable between individuals but it is generally accepted that this variant reduces enzyme activity by around 50% (PMID: 31337602). (SP) 1209 - This variant has been shown to be both maternally and paternally inherited (biallelic) (by trio analysis). (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign

It is observed in the gnomAD v2.1.1 dataset at total allele frequency of 3.184%. Functional studies provide strong evidence of the variant having a damaging effect on the gene or gene product (PMID: 10206677, 10206677, 9654207). In silico tool predictions suggest damaging effect of the variant on gene or gene product (REVEL: 0.77; 3Cnet: 0.88). Same nucleotide change resulting in same amino acid change has been previously reported as pathogenic/likely pathogenic with strong evidence (ClinVar ID: VCV000001900). The variant has been reported to be in trans with a pathogenic variant as either compound heterozygous or homozygous in at least 4 similarly affected unrelated individuals (PMID: 9654207). A different missense change at the same codon (p.Asp424Tyr) has been reported to be associated with BTD related disorder (PMID: 33312878). Therefore, this variant is classified as Pathogenic according to the recommendation of ACMG/AMP guideline.

This variant is also known as c.1270G>C (p.Asp424His) when using an alternative transcript (NM_001370658.1). This variant is a common cause of partial biotinidase deficiency (PMID: 10206677, 9654207, 12227467, 23644139); however, it has also been observed in individuals affected with profound biotinidase deficiency when this variant is in cis with another BTD variant, most commonly with the p.Ala171Thr variant, and in trans with a pathogenic variant (PMID: 10206677, 9654207). Functional characterization suggests that individuals who are homozygous for this variant typically have 50% of normal enzyme activity (PMID: 20539236, 28682309, 9654207), which is similar to the activity of unaffected heterozygotes of a profound biotinidase deficiency variant, and do not require biotin therapy (PMID: 20301497). However, homozygous individuals with activities in the partial deficiency range have been reported (PMID: 15060693). Untreated individuals with partial biotinidase deficiency are often asymptomatic in the absence of confounding factors such as significant illness (PMID: 9654207). It is present in the heterozygous state in the gnomAD population database at a frequency of 0.3% (9005/282830), including 199 homozygotes. In silico tools used to predict the effect of this variant on protein function yield discordant results. Analysis of the parental samples showed the mother is heterozygous and the father is heterozygous for this variant. Based on the available evidence, the c.1330G>C (p.Asp444His) variant is classified as Pathogenic.

ACMG classification criteria: PS3, PM3, PP3, PP4

This sequence change replaces aspartic acid, which is acidic and polar, with histidine, which is basic and polar, at codon 444 of the BTD protein (p.Asp444His). This variant is present in population databases (rs13078881, gnomAD 6%), and has an allele count higher than expected for a pathogenic variant. In the homozygous state this variant does not cause biotinidase deficiency or partial biotinidase deficiency (PMID: 28682309, 9654207). However, this variant in conjunction with another pathogenic variant is a common cause of partial biotinidase deficiency (PMID: 10206677, 9654207, 12227467, 23644139). This variant has also been observed in individuals affected with profound biotinidase deficiency when this variant is in cis with the p.A171T variant and in trans with a third variant (PMID: 10206677, 9654207). In individuals affected with partial biotinidase deficiency who harbor this variant in combination with another BTD variant, serum biotinidase activity was approximately 24% of the mean normal control activity (PMID: 9654207). In individuals affected with profound biotinidase deficiency who harbor this variant in cis with p.A171T and in trans with another BTD variant, serum biotinidase activity was <10% of the mean normal control activity (PMID: 10206677, 9654207). Individuals who are homozygous for this variant typically have an enzyme activity that is approximately 50% of normal (PMID: 20539236, 28682309, 9654207), similar to what is seen for a carrier of a profound allele. ClinVar contains an entry for this variant (Variation ID: 1900). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt BTD protein function with a positive predictive value of 95%. For these reasons, this variant has been classified as Pathogenic.

In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; Also known as c.1330 G>C, p.(D444H); This variant is associated with the following publications: (PMID: 24797656, 10206677, 29191167, 29359854, 36703223, 37772257, 37443404, 24525934, 11668630, 20981092, 21228398, 22975760, 22995991, 25087612, 25333069, 25795614, 24516753, 20532819, 17629531, 20556795, 9654207, 28498829, 27884173, 27535533, 27845546, 27657684, 29961769, 30912303, 30609409, 30487145, 31337602, 31028937, 31618753, 31980526, 34426522, 32746448, 33083013, 32235217, 35314707, 36007526, 31847883, 34738359, 35805799, 15060693, 20539236)

The observed invariant splice acceptor c.2042-2A>G variant in PIDD1 has been reported in individuals affected with PIDD1-related disorders (Teerlink CC, et. al., 2022). The c.2042-2A>G variant is reported with an allele frequency of 0.008% in the gnomAD Exomes. This variant has been submitted to the ClinVar database as Uncertain Significance / Likely Pathogenic. SpliceAI precits this variant to cause splice acceptor Loss (0.99). Loss of function variants have been previously reported to be disease causing. However, additional functional studies are required to prove the pathogenicity of the variant. For these reasons, this variant has been classified as Likely Pathogenic.

BTD: PM3:Very Strong, PM2:Supporting, PS3:Supporting

The BTD p.Asp444His variant is a well-known variant that causes partial Biotinidase (BTD) Deficiency (10-30% of normal enzyme activity) when found in the compound heterozygous state with a severe BTD mutation (Hymes_2001_PMID: 11668630). In addition, when the D444H variant is found on the same allele as A171T (double mutant), it is considered a pathogenic allele and can lead to profound BTD deficiency when found in the compound heterozygous state with another pathogenic allele (Norrgard_1998_PMID: 10206677). The variant was identified in dbSNP (ID: rs13078881), ClinVar (reported as pathogenic (11x), likely pathogenic (1x) and uncertain significance (1x)), Clinvitae and LOVD 3.0, but was not identified in the Cosmic or MutDB databases. The variant was identified in control databases in 9005 of 282830 chromosomes (199 homozygous) at a frequency of 0.031839 (Genome Aggregation Database Feb 27, 2017). The variant was observed in the following populations: European (Finnish) in 1396 of 25118 chromosomes (freq: 0.05558), European (non-Finnish) in 5119 of 129150 chromosomes (freq: 0.03964), South Asian in 1129 of 30616 chromosomes (freq: 0.03688), Ashkenazi Jewish in 340 of 10368 chromosomes (freq: 0.03279), Other in 217 of 7222 chromosomes (freq: 0.03005), Latino in 643 of 35436 chromosomes (freq: 0.01815), African in 159 of 24966 chromosomes (freq: 0.006369), and East Asian in 2 of 19954 chromosomes (freq: 0.0001). Liu et al. (2018) recently found that the D444H variant results in a decrease in protein expression with no loss of enzyme activity (Liu_2018_PMID: 29359854). Other studies have found 50% of normal BTD enzyme activity with one D444H allele present 25% activity with two D444H alleles present (Cowan_2010_PMID: 20539236), therefore indicating the effect of this variant on the protein. In summary, this variant is considered a hypomorphic allele and in combination with other variants in cis or trans, may result in reduced enzyme activity. Based on the above information this variant meets our laboratory’s criteria to be classified as pathogenic.

The BTD c.1330G>C variant is predicted to result in the amino acid substitution p.Asp444His. This sequence variant has been commonly documented to be causative for partial biotinidase deficiency (e.g., Norrgard et al. 1998. PubMed ID: 10206677; Swango et al. 1998. PubMed ID: 9654207; Wolf. 2012. PubMed ID: 22241090). Individuals homozygous for the c.1330G>C variant without an additional pathogenic variant are typically found to have ~50% of normal biotinidase enzyme activity, and are not usually clinically affected (Wolf. 2012, PubMed ID 22241090). Individuals compound heterozygous for the c.1330G>C variant and a second variant that causes profound biotinidase deficiency are expected to exhibit ~20-25% of enzyme activity, classified as partial biotinidase deficiency (Wolf. 2012. PubMed ID 22241090). This variant is reported in 5.6% of alleles in individuals of European (Finnish) descent in gnomAD. This variant is interpreted as pathogenic.

Variant interpretted as Pathogenic and reported on 12-12-2018 by Lab or GTR ID 26957. GenomeConnect assertions are reported exactly as they appear on the patient-provided report from the testing laboratory. GenomeConnect staff make no attempt to reinterpret the clinical significance of the variant.

Variant interpreted as Uncertain significance and reported on 02-05-2016 by Baylor. GenomeConnect-Invitae Patient Insights Network assertions are reported exactly as they appear on the patient-provided report from the testing laboratory. Registry team members make no attempt to reinterpret the clinical significance of the variant. Phenotypic details are available under supporting information.