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Functional analysis of MSH2 unclassified variants found in suspected Lynch syndrome patients reveals pathogenicity due to attenuated mismatch repair
  1. Eva AL Wielders1,
  2. Jan Hettinger1,2,
  3. Rob Dekker1,3,
  4. C Marleen Kets4,
  5. Marjolijn J Ligtenberg4,5,
  6. Arjen R Mensenkamp4,
  7. Ans MW van den Ouweland6,
  8. Judith Prins6,
  9. Anja Wagner6,
  10. Winand NM Dinjens7,
  11. Hendrikus Jan Dubbink7,
  12. Liselotte P van Hest8,
  13. Fred Menko8,
  14. Frans Hogervorst9,
  15. Senno Verhoef9,
  16. Hein te Riele1
  1. 1Division of Biological Stress Response, The Netherlands Cancer Institute, Amsterdam, The Netherlands
  2. 2Tumor Immunology Program, German Cancer Research Center (DKFZ), Immune Tolerance, Heidelberg, Germany
  3. 3RNA Biology and Applied Bioinformatics group, Swammerdam Institute for Life Sciences, University of Amsterdam, Amsterdam, The Netherlands
  4. 4Department of Human Genetics, Radboud University Medical Center, Nijmegen, The Netherlands
  5. 5Department of Pathology, Radboud University Medical Center, Nijmegen, The Netherlands
  6. 6Department of Clinical Genetics, Erasmus MC, University Medical Center, Rotterdam, The Netherlands
  7. 7Department of Pathology, Erasmus MC, University Medical Center, Rotterdam, The Netherlands
  8. 8Department of Clinical Genetics, VU University Medical Center, Amsterdam, The Netherlands
  9. 9The Netherlands Cancer Institute, Family Cancer Clinic, Amsterdam, The Netherlands
  1. Correspondence to Professor Hein te Riele, Division of Biological Stress Response, The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam 1066 CX, The Netherlands; h.t.riele{at}


Background Lynch syndrome, an autosomal-dominant disorder characterised by high colorectal and endometrial cancer risks, is caused by inherited mutations in DNA mismatch repair (MMR) genes. Mutations fully abrogating gene function are unambiguously disease causing. However, missense mutations often have unknown functional implications, hampering genetic counselling. We have applied a novel approach to study three MSH2 unclassified variants (UVs) found in Dutch families with suspected Lynch syndrome.

Methods The three mutations were recreated in the endogenous Msh2 gene in mouse embryonic stem cells by oligonucleotide-directed gene modification. The effect of the UVs on MMR activity was then tested using a set of functional assays interrogating the main MMR functions.

Results We recreated and functionally tested three MSH2 UVs: MSH2-Y165D (c.493T>G), MSH2-Q690E (c.2068C>G) and MSH2-M813V (c.2437A>G). We observed reduced levels of MSH2-Y165D and MSH2-Q690E but not MSH2-M813V proteins. MSH2-M813V was able to support all MMR functions similar to wild-type MSH2, whereas MSH2-Y165D and MSH2-Q690E showed partial defects.

Conclusions Based on the results from our functional assays, we conclude that the MSH2-M813V variant is not disease causing. The MSH2-Y165D and MSH2-Q690E variants affect MMR function and are therefore likely the underlying cause of familial cancer predisposition. Since the MMR defect is partial, these variants may represent low penetrance alleles.

  • Cancer: Colon
  • Genetic Screening/Counselling
  • Molecular Genetics
  • Lynch Syndrome
  • Mismatch Repair

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