Final Diagnosis
The Oncomine assay on the patient’s primary tumor reveals KRAS, PIK3CA, and CREBBP variants in addition to two MSH2 truncating variants with one at 49.9% and one at 35.6% variant allele frequency. Additionally, the tumor is found to have a TMB at the 82nd percentile of the MSKCC dataset and is MSI-High.
Discussion
The patient’s initial presentation of severe abdominal pain has a wide differential diagnosis and can include etiologies such as ectopic pregnancy, ovarian torsion, and appendicitis. However, given the strong family history of colorectal cancer, there is appropriate concern for Lynch syndrome. Lynch syndrome is an inherited autosomal dominant condition in which a mutation exists at germline frequency in the main MMR genes which include MLH1, PMS2, MSH2, and MSH6. Germline mutations in the EPCAM gene are also a cause of Lynch syndrome that result in epigenetic silencing of MSH2 and account for approximately 1-3% of cases. This condition pre-disposes patients to multiple cancers, particularly colorectal cancer, in young adulthood. In female patients, other malignancies include endometrial and ovarian cancers. Patients that subsequently develop malignancies often acquire a somatic mutation, or “second-hit,” in the second allele of the MMR gene that is affected by the germline mutation.
The MMR proteins function through dimerization and subsequent recognition and repair of mismatched bases and insertions or deletions, the latter particularly at microsatellite regions. These regions consist of 1-5 nucleotide repeats that span a length of about 10-60 base pairs and are interspersed throughout the genome. Deficiencies in the MMR proteins lead to instability of these regions, resulting in varying microsatellite lengths that can be analyzed through a PCR-based test as part of the Oncomine panel. This testing is recommended if patients meet clinical criteria based on their personal or family medical history.
The revised Bethesda criteria are a set of five main clinical guidelines to help distinguish those patients whose colorectal tumors should be tested for MSI. These include: a diagnosis of colorectal cancer under the age of 50, a second colorectal cancer or tumor associated with Lynch syndrome, histologic characteristics of an MSI-High tumor in a patient under the age of 60, a first-degree relative under the age of 50 with a Lynch syndrome-associated cancer, and two or more first- or second-degree relatives with a Lynch syndrome-associated cancer at any age. This patient met two of the criteria: the patient was less than 50 years of age at diagnosis with colorectal cancer and had two first or second-degree relatives that were diagnosed with a Lynch syndrome-associated cancer. Along with the recommended MSI testing for this patient, standard practice during the histologic assessment of colorectal tumors is to use immunohistochemical (IHC) stains for the four MMR proteins to visualize their level of expression.
As the IHC stains show, there is complete loss of MSH2 expression with reduced expression of its partner protein MSH6. The staining patterns of MLH1 and PMS2 show wild-type expression for both. This suggests that a mutation is present in MSH2 that results in an altered protein product that does not retain the IHC stain. Furthermore, as MSH2 and MSH6 function as a dimer, the IHC stain shows that MSH6 expression is reduced but not absent. This suggests that wild-type MSH6 expression requires wild-type MSH2 expression; therefore, if MSH2 is absent, the stability of MSH6 is markedly decreased.
Subsequently, the Oncomine assay shows that two MSH2 variants are detected with one truncating mutation at germline frequency and an additional truncating mutation at somatic frequency. This is suggestive of a “double-hit” of MSH2 whereby the somatic mutation is a secondary mutation and is the driver contributing to cancer development. The IHC results and the variants detected by Oncomine are therefore concordant and highlight a potential germline mutation in MSH2.
As evidenced by the TMB and MSI testing analysis by the Oncomine assay, the patient has a relatively high TMB when compared to the MSKCC dataset, and all tested MSI markers demonstrated instability. As a result of the testing, the patient is a candidate to have formal germline testing to confirm a diagnosis of Lynch syndrome. Of note, germline testing is performed on non-tumor tissue such as blood or saliva which can confirm if the patient harbors a mutant allele in one of the MMR genes. Furthermore, should a diagnosis of Lynch syndrome be rendered, the patient should receive the appropriate surveillance consisting of annual colonoscopy, upper endoscopy every 2-4 years, and endometrial sampling annually. In summary, this highlights the importance of molecular testing in cases of suspected Lynch syndrome to identify patients that require frequent screening for malignancies.
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