Whole-exome sequencing and RNA sequencing analyses of acinic cell carcinomas of the breast

Abstract

Aims: Acinic cell carcinoma (ACC) of the breast is a rare histological form of triple-negative breast cancer (TNBC). Despite its unique histology, targeted sequencing analysis has failed to identify recurrent genetic alterations other than those found in common forms of TNBC. Here we subjected three breast ACCs to whole-exome and RNA sequencing to determine whether they would harbour a pathognomonic genetic alteration.

Methods and results: DNA and RNA samples from three breast ACCs were subjected to whole-exome sequencing and RNA-sequencing, respectively. Somatic mutations, copy number alterations, mutational signatures and fusion genes were determined with state-of-the-art bioinformatics methods. Our analyses revealed TP53 hotspot mutations associated with loss of heterozygosity of the wild-type allele in two cases. Mutations affecting homologous recombination DNA repair-related genes were found in two cases, and an MLH1 pathogenic germline variant was found in one case. In addition, copy number analysis revealed the presence of a somatic BRCA1 homozygous deletion and focal amplification of 12q14.3-12q21.1, encompassing MDM2, HMGA2, FRS2, and PTPRB. No oncogenic in-frame fusion transcript was identified in the three breast ACCs analysed.

Conclusions: No pathognomonic genetic alterations were detected in the breast ACCs analysed. These tumours have somatic genetic alterations similar to those of common forms of TNBC, and may show homologous recombination deficiency or microsatellite instability. These findings provide further insights into why breast ACCs, which are usually clinically indolent, may evolve into or in parallel with high-grade TNBC.

Keywords: DNA damage repair; acinic cell carcinoma; breast cancer; massively parallel sequencing.

Figure 1.. Histologic characteristics of acinic cell carcinomas of the breast.

(A-B) Representative photomicrographs of hematoxylin and eosin (H&E)-stained acinic cell carcinomas of the breast (ACCs) from this study. (A-C) ACC1 displays an infiltrative growth (A) with microglandular features (B) and is composed of Paneth-like cells with coarse intracytoplasmic granules (C). (D-E) ACC12 displays a microglandular growth pattern (D) with cells featuring an amphophilic cytoplasm with fine granules (E). (F-H) ACC18 displays microglandular areas (F) composed of eosinophilic cells with coarse granules (G) and hypernephroid areas composed of clear cells (H). Scale bar, 200 μm (A, D), 50 μm (B, and E), 20 μm (C, G and H) and 100 μm (F).

Figure 2.. Repertoire of somatic mutations and mutational signatures of the acinic cell carcinomas of the breast.

(A) Non-synonymous somatic mutations affecting cancer-related genes^– and mutations shared among cases identified in the acinic cell carcinomas of the breast (ACCs; n=3) subjected to whole-exome sequencing (WES). Cases are shown in columns and genes in rows. (B) Copy number plots depicting segmented Log₂ ratios (y-axis) plotted according to genomic position (x-axis). Chromosomes are demarcated by alternating blue and gray colors (C) Mutational signatures of all somatic SNVs in breast ACCs (n=3). Pie charts indicate the proportion of the different mutational signatures identified in each case. (D) Representative hematoxylin and eosin micrograph of ACC12 arising in a patient with an MLH1 germline mutation (top) and micrograph depicting loss of MLH1 expression in the tumor cells (*). Normal breast (right lower corner) shows retention of MLH1 expression. Dashed line Scale bar, 50 μm. SNV, single nucleotide variant. Sig, signature; SNV, single nucleotide variant.