RESUMO
AIMS: The Lynch syndrome (LS) screening algorithm requires BRAF testing as a fundamental step to distinguish sporadic from LS-associated colorectal carcinomas (CRC). BRAF testing by immunohistochemistry (IHC) has shown variable results in the literature. Our aim was to analyse concordance between BRAFV600E IHC and BRAF molecular analysis in a large, mono-institutional CRC whole-slide, case series with laboratory validation. METHODS AND RESULTS: MisMatch repair (MMR) protein (hMLH1, hPMS2, hMSH2, and hMSH6) and BRAFV600E IHC were performed on all unselected cases of surgically resected CRCs (2018-2023). An in-house validation study for BRAFV600E IHC was performed in order to obtain optimal IHC stains. BRAFVV600E IHC was considered negative (score 0), positive (scores 2-3), and equivocal (score 1). Interobserver differences in BRAFV600E IHC scoring were noted in the first 150 cases prospectively collected. Nine-hundred and ninety CRCs cases (830 proficient (p)MMR/160 deficient (d)MMR) were included and all cases performed BRAFV600E IHC (BRAFV600E IHC-positive 13.5% of all series; 66.3% dMMR cases; 3.4% pMMR cases), while 333 also went to BRAF mutation analysis. Optimal agreement in IHC scoring between pathologists (P < 0.0001) was seen; concordance between BRAFV600E IHC and BRAF molecular analysis was extremely high (sensitivity 99.1%, specificity 99.5%; PPV 99.1%, and NPV 99.5%). Discordant cases were reevaluated; 1 score 3 + IHC/wildtype case was an interpretation error and one score 0 IHC/mutated case was related to heterogenous BRAFV600E IHC expression. Among the 12 IHC-equivocal score 1+ cases (which require BRAF molecular analysis), three were BRAF-mutated and nine BRAF-wildtype. CONCLUSION: BRAFV600E IHC can be used as a reliable surrogate of molecular testing after stringent in-house validation.
Assuntos
Neoplasias Encefálicas , Neoplasias Colorretais Hereditárias sem Polipose , Neoplasias Colorretais , Síndromes Neoplásicas Hereditárias , Humanos , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Neoplasias Colorretais Hereditárias sem Polipose/genética , Imuno-Histoquímica , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Detecção Precoce de Câncer , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Técnicas de Diagnóstico Molecular , Algoritmos , Reparo de Erro de Pareamento de DNA , MutaçãoRESUMO
BACKGROUND: Universal screening of colorectal cancer (CRC) patients for Lynch syndrome (LS) through MisMatch Repair (MMR) testing is recommended. BRAF V600E mutation and/or MLH1 promoter methylation (Reflex Testing, RefT)generally rule out LS in MLH1-deficient (dMLH1) patients. We estimated the impact of RefTon genetic counseling (GC) and on the diagnostic yield of genetic testing (GT). METHODS: Overall, 3199 CRC patients were referred to our center between 2011 and 2021. Patients referred until January 2019 (n=2536) underwent universal MMR testing and were termed 'Cohort A'; among patients after February 2019 (n=663), 'Cohort B', RefT was also performed in dMLH1 patients. RESULTS: Overall, 401/3199 patients (12.5%) were MMR-deficient (dMMR); 312 (77.8%) in cohort A and 89 (22.2%) inB; 346/401 were dMLH1 (86.3%), 262/312 (83.9%) in cohort A and 84/89 (94.3%) in B. In Cohort A, 91/312 (29.1%) dMMR patients were referred to GC, 69/91 (75.8%) were in the dMLH1 group; 57/69 (82.6%) dMLH1 patients underwent GT and 1/57 (1.7%) had LS. In Cohort B, 3/84 dMLH1 patients did not undergo BRAF testing. Three BRAF wt and not hypermethylated of the remaining 81 dMLH1 patients were referred to GC and GT, and one had LS. This diagnostic pathway reduced GC referrals by 96% (78/81) in Cohort B and increased the diagnostic yield of GT by about 20 times. CONCLUSION: Our findings support RefT in dMLH1 CRC patients within the LS diagnostic pathway, as it reduces the number of GC sessions needed and increases the diagnostic yield of GT.