title: A cell-free assay for the functional analysis of variants of the mismatch repair protein MLH1.
abstract: The hereditary colon and endometrium cancer predisposition Lynch Syndrome (also called HNPCC) is caused by a germ-line mutation in one of the DNA mismatch repair (MMR) genes. A significant fraction of the gene alterations detected in suspected Lynch Syndrome patients is comprised of amino acid substitutions. The relevance for cancer risk of these variants is difficult to assess, as currently no time- and cost-effective, validated, and widely applicable functional assays for the measurement of MMR activity are available. Here we describe a rapid, cell-free, and easily quantifiable MMR activity assay for the diagnostic assessment of variants of the MLH1 MMR protein. This assay allows the parallel generation and functional analysis of a series of variants of the MLH1 protein in vitro using readily available, or preprepared, reagents. Using this assay we have tested 26 MLH1 variants and of these, 15 had lost activity. These results are in concordance with those obtained from first-generation assays and with in silico and pathology data. After its multifocal technical and clinical validation this assay could have great impact for the diagnosis and counseling of carriers of an MLH1 variant and their relatives.
authors.affiliation: Departments of Toxicogenetics, Leiden University Medical Center, Leiden, The Netherlands.
authors.forename: Mark
authors.initials: M
authors.lastname: Drost
authors.forename: Jos e B M
authors.initials: Je
authors.lastname: Zonneveld
authors.forename: Linda
authors.initials: L
authors.lastname: van Dijk
authors.forename: Hans
authors.initials: H
authors.lastname: Morreau
authors.forename: Carli M
authors.initials: CM
authors.lastname: Tops
authors.forename: Hans F A
authors.initials: HF
authors.lastname: Vasen
authors.forename: Juul T
authors.initials: JT
authors.lastname: Wijnen
authors.forename: Niels
authors.initials: N
authors.lastname: de Wind
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics*; D002467:Cell Nucleus / Q000378:metabolism; D002474:Cell-Free System; D003001:Cloning, Molecular; D003123:Colorectal Neoplasms, Hereditary Nonpolyposis / Q000235:genetics*; D003362:Cost-Benefit Analysis; D004252:DNA Mutational Analysis / Q000379:methods; D004260:DNA Repair; D005816:Genetic Complementation Test; D014644:Genetic Variation; D006801:Humans; D008957:Models, Genetic; D000070957:MutL Protein Homolog 1; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods; D013997:Time Factors
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C501486:MLH1 protein, human; D009687:Nuclear Proteins; D000070957:MutL Protein Homolog 1