ClinVar Genomic variation as it relates to human health

The c.295C>T variant in APC is a missense variant predicted to cause substitution of Arginine by Tryptophan at amino acid position 99 (p.Arg99Trp). APC is defined by the ClinGen InSiGHT Hereditary Colorectal Cancer/Polyposis Variant Curation Expert Panel (HCCP VCEP) as a gene for which primarily truncating variants are known to cause disease (BP1). The highest population minor allele frequency of the variant c.295C>T in gnomAD v2.1.1 (non-cancer) is 0.0007355 (95/129170 alleles) in the European (non-Finnish) population, which is higher than the HCCP VCEP threshold (0.001%) for BS1 (BS1). This variant has been observed in heterozygous state in more than 1000 healthy unrelated adult individuals worth more than 10 healthy individual points in total (BS2; Invitae and Ambry Genetics internal data). It has also been observed once in a homozygous state (Ambry Genetics internal data). This variant has been observed 4 times with other APC variants deemed (likely) pathogenic by the HCCP VCEP in individuals with FAP (BP2; PMIDs 23159591, 25604157, Bonn internal data). Finally, this variant has been observed in 6 patients with an alternate molecular basis for disease (BP5; Leiden University Medical Center internal data). In summary, this variant meets the criteria to be classified as Benign for FAP based on the ACMG/AMP criteria applied, as specified by the HCCP VCEP: BS1, BS2, BP1, BP2 (VCEP specifications version 1; date of approval 12/12/2022).

This variant is considered likely benign or benign based on one or more of the following criteria: it is a conservative change, it occurs at a poorly conserved position in the protein, it is predicted to be benign by multiple in silico algorithms, and/or has population frequency not consistent with disease.

This variant is considered likely benign. This variant has been observed at a population frequency that is significantly greater than expected given the associated disease prevalence and penetrance.

This alteration is classified as likely benign based on a combination of the following: seen in unaffected individuals, population frequency, intact protein function, lack of segregation with disease, co-occurrence, RNA analysis, in silico models, amino acid conservation, lack of disease association in case-control studies, and/or the mechanism of disease or impacted region is inconsistent with a known cause of pathogenicity.

APC: BP1, BP2, BS3:Supporting, BS1

Variant identified in a genome or exome case(s) and assessed due to predicted null impact of the variant or pathogenic assertions in the literature or databases. Disclaimer: This variant has not undergone full assessment. The following are preliminary notes: Seen in one 38 year old with no polyps. Penetrance of APC is close to 100%. MaxMAF is 0.064% (higher than frequency of FAP).

Variant summary: The APC c.295C>T variant affects a non-conserved nucleotide, resulting in an amino acid change from a large size and basic Arg to a large size and aromatic Trp. 5/5 in-silico tools predict a damaging outcome for this variant, but these predictions have not been verified with functional studies. This variant was found in 48/121376 control chromosomes at a frequency of 0.0003955, which is about 7 times the maximal expected frequency of a pathogenic APC allele (0.0000602), suggesting this variant is benign. Furthermore, the variant was shown not to co-segregate with disease in one family (Dobbie_Eur J Cancer_1994), and the variant was reported to co-occur with deleterious APC variants, p.S1222X and large deletion of exon 15, respectively (Kerr_JMD_2013). In addition, several clinical laboratories classified this variant as benign/likely benign. Taken together, this variant was classified as benign.

The APC c.295C>T, p.Arg99Trp variant (rs139196838) has been reported in patients with attenuated familial adenomatous polyposis (Heinimann 2001, Jelsig 2016, Mindel 2011, Out 2015), but has also been found in trans with another APC pathogenic variant (Kerr 2013). The variant is listed in ClinVar (Variation ID: 135695), and observed in the general population at an overall frequency of 0.04% (115/277190 alleles) in the Genome Aggregation Database. The arginine at residue 99 is highly conserved, and computational algorithms (PolyPhen-2, SIFT) predict that the variant has an impact on the protein. Due to the limited information regarding this variant, its clinical significance could not be determined with certainty. References: Heinimann K et al. Nontruncating APC germ-line mutations and mismatch repair deficiency play a minor role in APC mutation-negative polyposis. Cancer Res. 2001; 61(20):7616-22. Jelsig A et al. Germline variants in Hamartomatous Polyposis Syndrome-associated genes from patients with one or few hamartomatous polyps. Scand J Gastroenterol. 2016; 51(9):1118-25. Kerr S et al. APC germline mutations in individuals being evaluated for familial adenomatous polyposis: a review of the Mayo Clinic experience with 1591 consecutive tests. J Mol Diagn. 2013; 15(1):31-43. Minde D et al. Messing up disorder: how do missense mutations in the tumor suppressor protein APC lead to cancer? Mol Cancer. 2011; 10:101. Out A et al. High-resolution melting (HRM) re-analysis of a polyposis patients cohort reveals previously undetected heterozygous and mosaic APC gene mutations. Fam Cancer. 2015; 14(2):247-57.

The APC p.Arg99Trp variant was identified in 7 of 3802 proband chromosomes (frequency: 0.002) from individuals or families with hamartomatous polyposis syndrome and familial adenomatous polyposis (Dobbie 1996, Heinimann 2001, Out 2015, Jelsig 2016, Kerr 2013). The variant was identified in dbSNP (rs139196838) as “with uncertain significance, other allele”, ClinVar (classified as uncertain significance by Counsyl, Mayo Clinic, Quest Diagnostics and 4 other submitters, likely benign by Color, Ambry Genetics, GeneDx and Sinai Health System and benign by Invitae and Integrated Genetics), LOVD 3.0 (observed 11x) and UMD-LSDB. The variant was identified in control databases in 113 of 282,846 chromosomes at a frequency of 0.0004 increasing the likelihood this could be a low frequency benign variant (Genome Aggregation Database Feb 27, 2017). The variant was observed in the following populations: European in 95 of 129,170 chromosomes (freq: 0.0007), African in 9 of 24,968 chromosomes (freq: 0.0004), Other in 2 of 7224 chromosomes (freq: 0.0003), Finnish in 3 of 25,118 chromosomes (freq: 0.0001), Latino in 3 of 35,436 chromosomes (freq: 0.00009), East Asian in 1 of 19,944 chromosomes (freq: 0.00005), while the variant was not observed in the Ashkenazi Jewish and South Asian populations. The variant was identified in individuals in trans with pathogenic APC variants (p.Ser1222* and exon 15 deletion) (Kerr 2013). The p.Arg99 residue is conserved in mammals and computational analyses (PolyPhen-2, SIFT, AlignGVGD, BLOSUM, MutationTaster) provide inconsistent predictions regarding the impact to the protein; this information is not very predictive of pathogenicity. The variant occurs outside of the splicing consensus sequence and 1 of 4 in silico or computational prediction software programs (SpliceSiteFinder, MaxEntScan, NNSPLICE, GeneSplicer) predict a greater than 10% difference in splicing; this is not very predictive of pathogenicity. In summary, based on the above information the clinical significance of this variant cannot be determined with certainty at this time although we would lean towards a more benign role for this variant. This variant is classified as likely benign.

This variant is classified as likely benign based on ACMG/AMP sequence variant interpretation guidelines (Richards et al. 2015 PMID: 25741868, with internal and published modifications).