BRAF testing modalities in histiocytic disorders: Comparative analysis and proposed testing algorithm

Objectives Understanding of histiocytic disorders has been revolutionized by demonstration of mitogen-activated protein kinase (MAPK) pathway mutations, most commonly BRAF(V600E). The optimal testing strategy to assess BRAF(V600E) is unknown. We aimed to compare performance of testing modalities, to propose a framework for evaluation of BRAF(V600E) mutation status in histiocytic disorders. Methods We retrospectively reviewed patients with histiocytic disorders and BRAF mutation testing on a lesional tissue specimen. Results In 120 patients, BRAF assessment included immunohistochemistry (IHC) in 97 (80.2%), polymerase chain reaction (PCR) in 35 (28.9%), and next-generation sequencing (NGS) in 62 (51.2%). Forty-five underwent both NGS and IHC. With NGS as the gold standard, the sensitivity and specificity of IHC were 82.4% and 96.4%. Three false negatives were observed in biopsy specimens with low BRAF(V600E) variant allele frequency or decalcified tissue. One false-positive IHC was observed in a lung biopsy specimen, likely due to antibody cross-reactivity with respiratory cilia. Among 14 with successful NGS and PCR, a single discordance was observed. Two PCR-to-IHC discrepancies were observed, including one other false-positive IHC. Conclusions Immunohistochemistry was highly specific for detection of BRAF(V600E). Main caveats were false negatives and lack of detection of non-BRAF(V600E) mutations. We propose the use of IHC as initial screening in general practice with reflex molecular testing if negative.