Germline Mutations in : Implications for Diagnostics in the ClinicMLH1: Implications for Diagnostics in the Clinic Leading to Isolated Loss of PMS2 Expression in Lynch Syndrome: Implications for Diagnostics in the Clinic
Lynch syndrome (LS) is characterized as familial clustering of colorectal and extracolonic tumors caused by deleterious mutations in one of the mismatch repair (MMR) genes MLH1, MSH2, MSH6, and PMS2. Germline mutations in MMR genes result in tumors that are marked by the presence of microsatellite instability and loss of expression of one or more MMR proteins, which are the tumor hallmarks of this disorder.1,2 The use of MMR immunohistochemistry (IHC) and microsatellite instability analysis in LS tumors are highly sensitive (77% to 89%) and specific (90%) in identifying patients with pathogenic mutations in MMR genes3 and, as a consequence, these methods have been largely used as first-line tests to guide genetic testing in LS suspected patients. Moreover, universal screening for LS using these methods, coupled with analysis of MLH1 promoter methylation and BRAF mutation for the detection of sporadic MMR deficiency, was recently proposed for all patients newly diagnosed with colorectal cancer (CRC)4 and is being adopted in major cancer centers.
The use of MMR IHC is very cost-effective as a guide to identify the most probably mutated MMR gene, reducing the cost and time burden of comprehensive genetic testing for the 4 LS genes. Functional DNA MMR activity requires the formation of protein heterodimers: MSH2 protein interacts with either MSH6 or MSH3, while MLH1 interacts either with PMS2 or PMS1 proteins. It is widely assumed that a functional defect in MLH1 results in the degradation of both MLH1 and PMS2, whereas a defect in PMS2 results only in the degradation of PMS2. Consequently, loss of expression of MLH1 and PMS2 generally indicates an alteration in MLH1 (somatic methylation of the MLH1 promoter region or MLH1 germline or somatic loss of function mutations) while solitary loss of PMS2 expression generally indicates an underlying germline defect in PMS2. The same effect is observed for the MSH2/MSH6 heterodimer, with the concomitant loss of MSH2 and MSH6 indicating a MSH2 germline mutation, and the isolated loss of MSH6 usually being caused by MSH6 mutations.
However, the correspondence of germline genetic mutation and loss of equivalent protein or its dimer is not observed in 100% of the cases.5–11 The most frequently reported discrepancy is the retention of MLH1 protein in germline MLH1 mutation carriers, usually accompanied by loss of PMS2 expression. This occurrence can be misleading for clinicians and genetic counselors, who are at risk of indicating the incorrect gene to be tested and, as a result, delaying a correct genetic diagnosis and proper patient management. In this letter, we comment on 2 recent papers, Dudley et al,9 and Rosty et al,10 that evaluate this phenomenon on a large cohorts with isolated loss of PMS2 expression and we also present the data of 4 Brazilian patients from our registry that presented this manifestation.
Since 1992, the Hereditary Colorectal Cancer Registry of AC Camargo Cancer Center has been recruiting families suspected to harbor CRC predisposing conditions. Selection criteria and detailed methods for the genetic investigations performed in this cohort were previously published.12,13 MMR IHC testing followed by germline mutation screening have been performed for a subset of these patients, who have consented to inform family history and participate in genetic testing (ethics committee approval number 870/06). In our cohort, we identified 4 families with an isolated loss of PMS2 immunostaining and germline pathogenic mutations in MLH1 instead of the expected mutations in PMS2. All families fulfilled the Amsterdam I or II clinical criteria. The identified MLH1 mutations include 2 splice site mutations in exon 7 and 10, 1 nonsense mutation in exon 17, and a frameshift mutation in exon 16 (Fig.