CMMRD-diagnostics in three cases: verification of PMS2 variants in homozygous status and investigation of their pathogenicity by cDNA-analyses

Monika Morak1,2, Melanie Locher2, Verena Steinke-Lange1,2, Nils Rahner3, Andreas Dufke4, Silke Redler3, Andreas Laner2, Udo Koehler2, Gabriela Moeslein5, Tanja Haeusser2, Trisari Massdorf1,2, Elke Holinski-Feder1,2

1Medizinische Klinik und Poliklinik IVI, Campus Innenstadt, Klinikum der Universität München, Munich, Germany. 2MGZ Medical Genetics Center, Munich, Germany. 3Medical Faculty, Institute of Human Genetics, Heinrich-Heine University, Düsseldorf, Germany. 4Institut für Medizinische Genetik und Angewandte Genomik, Universitätsklinikum Tübingen, Tübingen, Germany. 5Surgical Center for Hereditary Tumors, Helios University Hospital Wuppertal, Witten, Germany



The Constitutional Mismatch Repair Deficiency (CMMRD) is a hereditary cancer predisposition for hematological, brain, and other tumors starting from childhood caused by biallelic variants in MLH1, MSH2, MSH6, or PMS2. PMS2-analysis is error-prone due to the presence of multiple pseudogenes. Therefore, in three patients (P1, P2, P3) with clinical suspicion of CMMRD, the homozygous status of PMS2 variants and their pathogenicity were verified by additional cDNA- and MLPA-analyses.


PMS2 germline-analyses involve NGS or Sanger sequencing of Long-Range PCRs, MLPA and a specific SVA PCR. RNA was isolated from patient blood-lymphocytes cultured for short-term in absence/presence of puromycin (cDNA-P/+P) to investigate transcripts with active/inhibited nonsense-mediated mRNA decay (NMD). From cDNAs, the full-length PMS2 transcript was amplified and sequenced.


In P1 with a homozyogous PMS2 exon 12 duplication, cDNA-analysis showed exon 12 duplication r.2007_2174dup p.Lys670_Ala725dup in the transcript in absence of a wildtype allele. The segregation analysis found both parents heterozygous for the duplication of PMS2 exon 12.

In P2, a SINE-VNTR-Alu-insertion in intron 7 resulted in the insertion r.803_804insAATGTGCCATGTGAACCACCCCGTCTGAAAAGTGAGGAGCCCCTCTGCCCGGCAGCCGCCCCGTCTGGGAG p.Tyr268* in cDNA+P in absence of a wildtype allele. In cDNA-P, the amplification failed, indicating that both alleles were mutated and transcripts were subjected to NMD.

For P3 with variant c.736_741delinsTGTGTGTGAAG p.(Pro246Cysfs*3) identified homozygously by Sanger sequencing, no cDNA was available, but in MLPA-analysis a complete dropout of the probe hybridising to the variants` position verified its homozygous status.


By performing cDNA-analysis or MLPA we confirm presence and pathogenicity of homozygous PMS2 variants in three CMMRD-patients.

Abstract references

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