Variant classification summary

NM_000249.4:c.1682A>G

MLH1 · NP_000240.1:p.(Tyr561Cys) · NM_000249.4

GRCh37: chr3:37083773 A>G · GRCh38: chr3:37042282 A>G

Gene: MLH1 Transcript: NM_000249.4

Final call

VUS

PM2 supporting

Variant details

Gene

MLH1

Transcript

NM_000249.4

Protein

NP_000240.1:p.(Tyr561Cys)

gnomAD AF

ClinVar

OncoKB

Unknown Oncogenic Effect

Classification rationale

Interpretation summary

Generated evidence synthesis

1

The MLH1 c.1682A>G (p.Tyr561Cys; p.Y561C) variant has not been observed in somatic cancers in COSMIC and has been reported in ClinVar with uncertain significance submissions.

2

This variant is present at very low frequency in population databases, with gnomAD v4.1 total allele frequency 3.10936e-06 and grpmax filtering allele frequency 8e-07, which is below the MLH1 PM2_Supporting threshold of 0.00002.

3

No variant-specific calibrated functional assay result for this variant was identified in the reviewed MMR functional assay materials, so functional evidence was not used to support or refute pathogenicity.

4

SpliceAI predicts no significant splice impact for this variant, with a maximum delta score of 0.01, and available in silico evidence was insufficient to assign the MLH1 missense PP3 or BP4 rules without an HCI-prior value.

Final determination: No criteria-combination rule matched the adjudicated criteria in the ClinGen InSiGHT Hereditary Colorectal Cancer/Polyposis Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for MLH1 Version 2.0.0 v2.0.0 framework, so the variant remains a Variant of Uncertain Significance pending human review.

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
BA1	Not met	Population frequency does not meet the MLH1 BA1 threshold. In gnomAD v4.1, the highest filtering allele frequency is 8e-07, which is below the BA1 threshold of 0.001 (0.1%).	cspec gnomad_v4
BS1	Not met	Population frequency does not meet the MLH1 BS1 threshold. In gnomAD v4.1, the highest filtering allele frequency is 8e-07, which is below the BS1 range of 0.0001 to less than 0.001.	cspec gnomad_v4
BS2	Not assessed	No confirmed observation of this variant in trans with a known pathogenic MLH1 variant in an individual without clinical evidence of constitutional mismatch repair deficiency was identified.	cspec vcep_t_a_b_l_e___f_o_r___c_m_m_r_d___d_i_a_g_n_o_s_i_s
BS3	Not assessed	No variant-specific calibrated functional evidence showing proficient MLH1 function was identified, so BS3 cannot be applied.	cspec vcep_f_u_n_c_t_i_o_n_a_l___a_s_s_a_y___s_v_i___d_o_c_u_m_e_n_t_a_t_i_o_n___m_m_r vcep_f_u_n_c_t_i_o_n_a_l___a_s_s_a_y___f_l_o_w_c_h_a_r_t
BS4	Not assessed	No family segregation study showing lack of co-segregation with Lynch syndrome-associated disease was identified.	cspec
BP1	N/A	BP1 is not applicable for this VCEP framework.	cspec
BP2	N/A	BP2 is not applicable for this VCEP framework.	cspec
BP3	N/A	BP3 is not applicable for this VCEP framework.	cspec
BP4	Not assessed	SpliceAI predicts no significant splice impact for this variant with a maximum delta score of 0.01, which is below the non-canonical splice threshold of 0.1; however, this is a missense variant, and the MLH1 BP4 missense rule requires an HCI-prior probability below 0.11, which was not available.	cspec spliceai
BP5	Not assessed	No tumor data showing mismatch repair findings inconsistent with MLH1-associated disease, and no qualifying MLH1 methylation or BRAF V600E evidence, were identified.	cspec
BP6	N/A	BP6 is not applicable for this VCEP framework.	cspec
BP7	N/A	BP7 is a silent or intronic variant rule and does not apply to this missense variant.	cspec
PM1	N/A	PM1 is not applicable for this VCEP framework because recognized mutational hot spots are not used for MMR variant classification.	cspec hotspots
PM2	Met	Population frequency is below the MLH1 PM2_Supporting threshold. In gnomAD v4.1, the total allele frequency is 3.10936e-06 (5/1608050 alleles) with grpmax filtering allele frequency 8e-07, which is below the threshold of 0.00002 (<1 in 50,000 alleles).	cspec gnomad_v4 gnomad_v2
PM3	Not assessed	No confirmed in trans observation with another pathogenic or likely pathogenic MLH1 variant in a patient meeting constitutional mismatch repair deficiency criteria was identified.	cspec vcep_t_a_b_l_e___f_o_r___c_m_m_r_d___d_i_a_g_n_o_s_i_s
PM4	N/A	PM4 is not applicable for this VCEP framework.	cspec
PM5	Not assessed	No previously established pathogenic or likely pathogenic missense comparator at MLH1 codon 561 was identified in the reviewed VCEP materials, and PP3 was not assigned; therefore PM5 was not applied.	cspec vcep_v_c_e_p___p_i_l_o_t___v_a_r_i_a_n_t_s_______m_m_r
PM6	N/A	PM6 is not applicable for this VCEP framework.	cspec
PP1	Not assessed	No segregation data showing co-segregation of this variant with Lynch syndrome-associated disease in affected relatives were identified.	cspec
PP2	N/A	PP2 is not applicable for this VCEP framework.	cspec
PP3	Not assessed	SpliceAI does not support a splice defect for this variant, with a maximum delta score of 0.01, and the MLH1 PP3 missense rule requires an HCI-prior probability above 0.68 or 0.81, which was not available. Available in silico evidence is therefore insufficient to apply PP3.	cspec spliceai
PP4	Not assessed	No tumor microsatellite instability, immunohistochemistry, or MLH1 promoter methylation data were identified to support an MLH1-consistent mismatch repair-deficient phenotype.	cspec
PP5	N/A	PP5 is not applicable for this VCEP framework.	cspec
PS1	Not assessed	No previously established MLH1 pathogenic or likely pathogenic variant encoding the same amino acid change by a different nucleotide change was identified in the reviewed VCEP materials, so PS1 was not applied.	cspec vcep_v_c_e_p___p_i_l_o_t___v_a_r_i_a_n_t_s_______m_m_r
PS2	Not assessed	No confirmed de novo occurrence of this variant in an individual with an MLH1-consistent mismatch repair-deficient tumor was identified.	cspec
PS3	Not assessed	No variant-specific calibrated functional assay result demonstrating damaging MLH1 function was identified, so PS3 cannot be applied.	cspec vcep_f_u_n_c_t_i_o_n_a_l___a_s_s_a_y___s_v_i___d_o_c_u_m_e_n_t_a_t_i_o_n___m_m_r vcep_f_u_n_c_t_i_o_n_a_l___a_s_s_a_y___f_l_o_w_c_h_a_r_t
PS4	N/A	PS4 is not applicable for this VCEP framework because proband counting is not used for MMR variant classification when tumor immunohistochemistry data are available for PP4.	cspec clinvar
PVS1	Not met	This variant is a missense substitution, p.Tyr561Cys, and does not fall into the MLH1 PVS1 null-variant categories. The variant-level PVS1 assessment states that it is not a nonsense, frameshift, or canonical ±1/2 splice variant, so PVS1 is not applied.	cspec pvs1_gene_context pvs1_variant_assessment

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