title: Genetic screens to identify pathogenic gene variants in the common cancer predisposition Lynch syndrome.
abstract: In many individuals suspected of the common cancer predisposition Lynch syndrome, variants of unclear significance (VUS), rather than an obviously pathogenic mutations, are identified in one of the DNA mismatch repair (MMR) genes. The uncertainty of whether such VUS inactivate MMR, and therefore are pathogenic, precludes targeted healthcare for both carriers and their relatives. To facilitate the identification of pathogenic VUS, we have developed an in cellulo genetic screen-based procedure for the large-scale mutagenization, identification, and cataloging of residues of MMR genes critical for MMR gene function. When a residue identified as mutated in an individual suspected of Lynch syndrome is listed as critical in such a reverse diagnosis catalog, there is a high probability that the corresponding human VUS is pathogenic. To investigate the applicability of this approach, we have generated and validated a prototypic reverse diagnosis catalog for the MMR gene MutS Homolog 2 (Msh2) by mutagenizing, identifying, and cataloging 26 deleterious mutations in 23 amino acids. Extensive in vivo and in vitro analysis of mutants listed in the catalog revealed both recessive and dominant-negative phenotypes. Nearly half of these critical residues match with VUS previously identified in individuals suspected of Lynch syndrome. This aids in the assignment of pathogenicity to these human VUS and validates the approach described here as a diagnostic tool. In a wider perspective, this work provides a model for the translation of personalized genomics into targeted healthcare.
authors.affiliation: Department of Toxicogenetics, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
authors.forename: Mark
authors.initials: M
authors.lastname: Drost
authors.forename: Anne
authors.initials: A
authors.lastname: Lützen
authors.forename: Sandrine
authors.initials: S
authors.lastname: van Hees
authors.forename: Daniel
authors.initials: D
authors.lastname: Ferreira
authors.forename: Fabienne
authors.initials: F
authors.lastname: Calléja
authors.forename: José B M
authors.initials: JB
authors.lastname: Zonneveld
authors.forename: Finn Cilius
authors.initials: FC
authors.lastname: Nielsen
authors.forename: Lene Juel
authors.initials: LJ
authors.lastname: Rasmussen
authors.forename: Niels
authors.initials: N
authors.lastname: de Wind
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D001483:Base Sequence; D003123:Colorectal Neoplasms, Hereditary Nonpolyposis / Q000235:genetics*; D020022:Genetic Predisposition to Disease / Q000235:genetics*; D005820:Genetic Testing; D014644:Genetic Variation; D006801:Humans; D018895:Microsatellite Repeats / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D051718:MutS Homolog 2 Protein / Q000737:chemistry / Q000172:deficiency / Q000235:genetics*; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics; D059386:Reverse Genetics / Q000379:methods; D017422:Sequence Analysis, DNA; D013866:Thioguanine; D014774:Virulence / Q000235:genetics
chemical_list: C497172:MSH2 protein, human; D051718:MutS Homolog 2 Protein; D013866:Thioguanine