Finding clues to fibroepithelial tumours using next generation sequencing

How do breast tumours become what they are? We investigated what could potentially lead to this group of fibroepithelial lesions (FELs), comprising fibroadenoma (FA) and phyllodes tumours (PT). FAs are more common compared to PT, but at times it is challenging to distinguish them based on how they look. PTs tend to be more clinically aggressive, and its malignant subtype could even metastasize and may be fatal, thus making it important for us to search for better diagnostic and prognostic methods.

Targeted next generation sequencing (NGS) offers a viable option in profiling genomic DNA from formalin-fixed paraffin-embedded (FFPE) material. Adjacent normal breast FFPE is often limited and more challenging to acquire, making it less feasible for platforms such as whole exome sequencing and whole genome sequencing which require both tumour and normal DNA, and a higher amount of sample input. We performed NGS on an international cohort of these tumours [1] with a custom amplicon-based panel that targets 16 genes. This gene set was established based on our previous studies [2,3], covering MED12, TERT promoter, SETD2, KMT2D, RARA, FLNA, NF1, PIK3CA, EGFR, RB1, BCOR, TP53, PTEN, ERBB4, IGF1R, and MAP3K1. The Illumina HiSeq 4000 was used to sequence 150 bp paired-end reads at a depth of at least 100x.

Figure 1. Genomic progression model for MED12-related fibroepithelial lesions

What we found were frequent MED12 mutations in both FAs and PTs. The latter had a higher tendency of harbouring MED12, TERT promoter, RARA, FLNA, SETD2, KMT2D, and MAP3K1 than FAs, suggesting their role in progression from FAs to PTs. We noted additional gene alterations in borderline and malignant PTs than benign PTs in ERBB4, PIK3CA, RB1, TP53 and NF1, which likely play a part in malignant transformation [2,3]. There were significantly higher rates of TERT promoter, RARA, SETD2, EGFR, ERBB4, MAP3K1, and IGF1R aberrations in FELs with mutant MED12. However, FELs with wild-type MED12 were more likely to possess TP53 and PIK3CA mutations [1]. Our study affirms the role of a set of genes in FELs, including its utility in refining PT diagnosis [4], predicting PT in core biopsies [5], and differentiating malignant PT from spindle cell metaplastic breast carcinoma [6] and breast sarcomas [7].

References 1. Md Nasir ND, Ng CCY, Rajasegaran V, et al. Genomic characterisation of breast fibroepithelial lesions in an international cohort. J Pathol. 2019 Dec;249(4):447-460. 2. Lim WK, Ong CK, Tan J, et al. Exome sequencing identifies highly recurrent MED12 somatic mutations in breast fibroadenoma. Nat Genet. 2014 Aug;46(8):877-80. 3. Tan J, Ong CK, Lim WK, et al. Genomic landscapes of breast fibroepithelial tumors. Nat Genet. 2015 Nov;47(11):1341-5. 4. Koh VCY, Ng CCY, Bay BH, et al. The utility of a targeted gene mutation panel in refining the diagnosis of breast phyllodes tumours. Pathology 2019;51:531-534. 5. Sim Y, Ng GXP, Ng CCY, et al. A novel genomic panel as an adjunctive diagnostic tool for the characterization and profiling of breast Fibroepithelial lesions. BMC Med Genomics 2019;12:142. 6. Yeong J, Thike AA, Young Ng CC, et al. A genetic mutation panel for differentiating malignant phyllodes tumour from metaplastic breast carcinoma. Pathology 2017;49:786-789. 7. Lim SZ, Ng CCY, Rajasegaran V, et al. Genomic profile of breast sarcomas: a comparison with malignant phyllodes tumours. Breast Cancer Res Treat 2019;174:365-373.

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Ms Nur Diyana Md Nasir

Research Officer, Department of Anatomical Pathology

Singapore General Hospital