ClinVar Genomic variation as it relates to human health

Variant summary: CYP27A1 c.1184+1G>A is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Three predict the variant abolishes the canonical 5' splicing donor site. At least one publication reports experimental evidence that this variant affects mRNA splicing by skipping of exon 7 and joining exon 6 to exon 8 that translates to a sequence of 28 novel amino acids preceding a termination codon (Garuti_1996). The variant allele was found at a frequency of 0.00016 in 251280 control chromosomes. This frequency is not significantly higher than estimated for a pathogenic variant in CYP27A1 causing Cerebrotendinous Xanthomatosis (0.00016 vs 0.0011), allowing no conclusion about variant significance. c.1184+1G>A has been reported in the literature as homozygous and compound heterozygous genotypes in multiple individuals affected with Cerebrotendinous Xanthomatosis (example, Garuti_1996, Gianneschi_2013). These data indicate that the variant is very likely to be associated with disease. Eight clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All laboratories classified the variant as pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic.

NM_000784.3(CYP27A1):c.1184+1G>A is a canonical splice variant classified as pathogenic in the context of cerebrotendinous xanthomatosis. c.1184+1G>A has been observed in cases with relevant disease (PMID: 22878431). Functional assessments of this variant are available in the literature (PMID: 9392430). c.1184+1G>A has been observed in population frequency databases (gnomAD: SAS 0.07%). In summary, NM_000784.3(CYP27A1):c.1184+1G>A is a canonical splice variant in a gene where loss of function is a known mechanism of disease, is predicted to disrupt protein function, and has been observed more frequently in cases with the relevant disease than in healthy populations. Please note: this variant was assessed in the context of healthy population screening.

PP3, PM3, PS3, PS4, PVS1

The c.1184+1G>A intronic alteration consists of a G to A substitution one nucleotide after coding exon 6 of the CYP27A1 gene. Alterations that disrupt the canonical splice donor site are typically deleterious in nature (Richards, 2015). Based on data from gnomAD, the A allele has an overall frequency of 0.02% (46/282682) total alleles studied. The highest observed frequency was 0.07% (22/30614) of South Asian alleles. This alteration has been detected in the homozygous state and in trans with other disease-causing CYP27A1 alterations in multiple unrelated individuals diagnosed with cerebrotendinous xanthomatosis (Stelten, 2018; Lipiski, 2020; Lipiski, 2020; Rashvand, 2021; Ginanneschi, 2013; Lee, 2001). This nucleotide position is highly conserved in available vertebrate species. Experimental studies show this alteration results in abnormal splicing and introduces a frameshift (Garuti, 1997). In silico splice site analysis predicts that this alteration will weaken the native splice donor site. Based on the available evidence, this alteration is classified as pathogenic.

Observed in the homozygous state or with a pathogenic variant on the opposite allele (in trans) in patients with cerebrotendinous xanthomatosis referred for genetic testing at GeneDx and in published literature (Garuti et al., 1997; Verrips et al., 2000; Lee et al., 2001; Lipinski et al., 2020); Canonical splice site variant and published functional studies demonstrate that this variant leads to the production of three abnormal transcripts, and almost undetectable levels of mRNA in patient fibroblasts (Garuti et al., 1997); This variant is associated with the following publications: (PMID: 10775536, 25525159, 11181744, 26643207, 20402754, 28894950, 29260356, 32793533, 33659184, 34066437, 33400472, 31589614, 33083013, 33520900, 9392430)

This sequence change affects a donor splice site in intron 6 of the CYP27A1 gene. RNA analysis indicates that disruption of this splice site induces altered splicing and may result in an absent or disrupted protein product. This variant is present in population databases (rs587778777, gnomAD 0.07%). Disruption of this splice site has been observed in individuals with cerebrotendinous xanthomatosis (PMID: 9392430, 21645175, 28894950). This variant is also known as In6+1G>A. ClinVar contains an entry for this variant (Variation ID: 65833). Studies have shown that disruption of this splice site results in skipping of 89 nucleotides of exon 6 and introduces a premature termination codon (PMID: 9392430). The resulting mRNA is expected to undergo nonsense-mediated decay. For these reasons, this variant has been classified as Pathogenic.

The observed invariant splice acceptor variant c.1184+1G>A in CYP27A1 gene has been reported previously in both homozygous and compound heterozygous state in multiple inviduals affected with cerebrotendinous xanthomatosis (Ginanneschi et al. 2013; Rashvand et al. 2021). Experimental evidence shows that this variant activates a cryptic donor splice site and leads to skipping of 89 base pairs of exon 6, introduces a new reading frame and creates a premature stop codon that does not three essential amino acids (Lys354, Lys358, and Arg362) that are important for the ferredoxin-binding domain (Rashvand et al. 2021). The c.1184+1G>A variant is present with an allele frequency of 0.02% on gnomAD Exomes database. This variant has been submitted to the ClinVar database as Likely Pathogenic / Pathogenic (multiple submissions). SpliceAI predicts a splice donor loss of 0.99 for this variant. Loss of function variants in CYP27A1 gene have been previously reported to be disease causing. For these reasons, this variant has been classified as Pathogenic. In the absence of another reportable variant in the CYP27A1 gene, the molecular diagnosis is not confirmed.

Converted during submission to Pathogenic.

The CYP27A1 c.1184+1G>A variant is predicted to disrupt the GT donor site and interfere with normal splicing. This variant has been reported in the homozygous and compound heterozygous states in individuals with cerebrotendinous xanthomatosis (see for example Garuti et al. 1997. PubMed ID: 9392430; Lipiński et al. 2020. PubMed ID: 32793533). This variant is reported in 0.072% of alleles in individuals of South Asian descent in gnomAD. Variants that disrupt the consensus splice donor site in CYP27A1 are expected to be pathogenic, and this variant has been classified as pathogenic by multiple independent submitters to the ClinVar database (https://www.ncbi.nlm.nih.gov/clinvar/variation/65833). Given all the evidence, we interpret c.1184+1G>A as pathogenic.