Variant: NC_012920.1(MT-CYB):m.3243A>T

CA913168968

689856 (ClinVar)

Gene: MT-TL1

Condition: mitochondrial disease

Inheritance Mode: Mitochondrial inheritance

UUID: 63fd61ad-6e82-43d1-952a-16c083e6ec90

Approved on: 2022-10-10

Published on: 2023-03-13

HGVS expressions

NC_012920.1:m.3243A>T
J01415.2:m.3243A>T

Likely Pathogenic

The Expert Panel has overridden the computationally generated classification - "Uncertain Significance - Insufficient Evidence"

• Met criteria codes: PM5_Supporting PS3_Moderate PS4_Moderate PM2_Supporting PP3

• Not Met criteria codes: PM6 PS2 PP1

Expert Panel

Mitochondrial Diseases VCEP

Criteria Specification Information

Criteria Specification: ClinGen Mitochondrial Disease Nuclear and Mitochondrial Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines Version 1_mtDNA

Evidence submitted by expert panel

Mitochondrial Diseases VCEP

The m.3243A>T variant in MT-TL1 has been reported in eight unrelated probands with primary mitochondrial disease. Features in affected individuals include mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) and chronic progressive external ophthalmoplegia (CPEO), as well as myoclonic epilepsy, exercise intolerance with rhabdomyolysis, cataracts, sensorineural hearing loss, diabetes, peripheral neuropathy, and ataxia (PMIDs: 9168904, 18203188, 20471262, 23220830, 30210801, 33924034, 25504047; PS4_moderate). In all individuals, the variant was present at heteroplasmy, ranging from 69-99% in muscle and 13-50% in blood. Age of onset varied from six to 29 years old. There are no large families reported in the medical literature to consider for evidence of segregation. There are no de novo occurrences of this variant reported to our knowledge. The m.3243A>T variant is absent in the GenBank dataset and gnomAD3.1.2. There is one heteroplasmic occurrence in 195,893 sequences in the Helix database, however the overall frequency is still low (PM2_supporting). The computational predictor MitoTIP suggests this variant is pathogenic, scoring in the 58.8 percentile, and HmtVAR also predicts it to be pathogenic with a score of 0.8 (PP3). The m.3243A>T variant is at the same nucleotide position as a previously established pathogenic variant (m. 3243A>G; PM5_supporting). Several different functional studies have been performed on this variant that support its pathogenicity and show independent deleterious effects of the variant, including generation of cybrids, single fiber testing, and aminoacylation assays (PS3_moderate). Cybrids with high levels of this variant, when compared to wild type, showed lower oxygen consumption, higher glucose consumption and lactate production, increased motility and migration capacities associated with specific glycosylation patterns, lower growth rate, and altered cellular processes (PMIDs: 25504047, 32273537). Single fiber testing showed higher levels of the variant in COX-deficient fibers (95.8 ± 0.7%, n=9) than in COX-positive fibers (44.3 ± 8.8%, n=9; p < 0.001; PMID: 20471262). Structural and aminoacylation studies of in-vitro transcribed human tRNALeu(UUR) showed that the m.3243A>T variant was associated with a 300-fold loss of aminoacylation efficiency (PMID: 12729737). In summary, this variant meets criteria to be classified as likely pathogenic for primary mitochondrial disease inherited in a mitochondrial manner. We note that one expert on this panel felt pathogenic was a more appropriate classification given the functional evidence of impact however the majority agreed with likely pathogenic. This classification was approved by the NICHD/NINDS U24 ClinGen Mitochondrial Disease Variant Curation Expert Panel on October 10, 2022. Mitochondrial DNA-specific ACMG/AMP criteria applied (PMID: 32906214): PS4_moderate, PM2_supporting, PP3, PM5_supporting, PS3_moderate.

Met criteria codes

• PM5_Supporting: Same nucleotide position as previously established pathogenic variant m.3243A>G.
• PS3_Moderate: The m.3243A>T variant is at the same nucleotide position as a previously established pathogenic variant (m. 3243A>G; PM5_supporting). Several different functional studies have been performed on this variant that support its pathogenicity and show independent deleterious effects of the variant, including generation of cybrids, single fiber testing, and aminoacylation assays (PS3_moderate). Cybrids with high levels of this variant, when compared to wild type, showed lower oxygen consumption, higher glucose consumption and lactate production, increased motility and migration capacities associated with specific glycosylation patterns, lower growth rate, and altered cellular processes (PMIDs: 25504047, 32273537). Single fiber testing showed higher levels of the variant in COX-deficient fibers (95.8 ± 0.7%, n=9) than in COX-positive fibers (44.3 ± 8.8%, n=9; p < 0.001; PMID: 20471262). Structural and aminoacylation studies of in-vitro transcribed human tRNALeu(UUR) showed that the m.3243A>T variant was associated with a 300-fold loss of aminoacylation efficiency (PMID: 12729737).
• PS4_Moderate: The m.3243A>T variant in MT-TL1 has been reported in eight unrelated probands with primary mitochondrial disease. Features in affected individuals include mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) and chronic progressive external ophthalmoplegia (CPEO), as well as myoclonic epilepsy, exercise intolerance with rhabdomyolysis, cataracts, sensorineural hearing loss, diabetes, peripheral neuropathy, and ataxia (PMIDs: 9168904, 18203188, 20471262, 23220830, 30210801, 33924034, 25504047; PS4_moderate). In all individuals, the variant was present at heteroplasmy, ranging from 69-99% in muscle and 13-50% in blood. Age of onset varied from six to 29 years old.
• PM2_Supporting: The m.3243A>T variant is absent in the GenBank dataset and gnomAD3.1.2. There is one heteroplasmic occurrence in 195,893 sequences in the Helix database, however the overall frequency is still low (PM2_supporting).
• PP3: The computational predictor MitoTIP suggests this variant is pathogenic, scoring in the 58.8 percentile, and HmtVAR also predicts it to be pathogenic with a score of 0.8 (PP3).

Not Met criteria codes

• PM6: There are no de novo occurrences of this variant reported to our knowledge.
• PS2: There are no de novo occurrences of this variant reported to our knowledge.
• PP1: There are no large families reported in the medical literature to consider for evidence of segregation.