Genetic Analysis of Early Neoplasia in the Breast: Next-Generation Sequencing of Flat Epithelial Atypia and Associated Ductal and Lobular Lesions

The molecular features of invasive breast cancers (IBC) have been well-characterized, but less is known about the earlier stages of neoplasia, including oncogenic drivers in early intraductal lesions. Flat epithelial atypia (FEA) is considered the earliest recognized precursor in the low-grade neoplasia pathway, but its mutational repertoire has not been studied, and drivers of the transition to morphologically more advanced lesions are unknown. Herein, we utilized next-generation sequencing to analyze 39 synchronous lesions from 13 patients, including FEA (n = 12) or predominantly FEA with early atypical ductal hyperplasia (FEA/early atypical ductal hyperplasia [ADH], n = 5) and associated ADH (n = 2), ductal carcinoma in situ (ductal carcinoma in situ [DCIS], n = 11), lobular carcinoma in situ (n = 3), and/or IBC with ductal and/or lobular differentiation (n = 6). Aside from 1 DCIS sample, all sequenced lesions in each patient were clonally related to one another. Recurrent alterations in FEA and FEA/early ADH included PIK3CA (69%), NCOR1 (31%), CBFB (31%), RUNX1 (15%), and GATA3 (23%). The mutational repertoire of FEA was similar to The Cancer Genome Atlas luminal IBC, except CBFB and NCOR1 mutations, which were more frequent in FEA and (along with PIK3CA, FOXA1, and CDKN1B) not always identified in paired morphologically advanced lesions. Compared with FEA, DCIS had more mutations and chromosomal copy number changes, including aberrations in PI-3 kinase pathway, transcription factors, chromatin remodeling genes, and TP53. CDH1 mutations identified in lobular carcinoma in situ were absent in paired FEA. Analysis of cases with ductal and lobular heterogeneity, including Rosen's triad, confirmed the shared clonality of the ductal and lobular components with features of genetic divergence. IBC of no special type were genetically similar to DCIS, and tubular carcinomas were similar to FEA. The results reveal the mutational repertoire of FEA and the genetics of early breast neoplasia, highlighting the clonal relationships of FEA to ductal and lobular carcinomas. Luminal breast cancer-associated genetic alterations are present at the earliest morphologically recognized stages of neoplasia.