Introduction: Invasive lobular carcinoma (ILC) is the most prevalent histologic special type of breast cancer, representing ~15% of invasive breast cancers. ILCs are mostly hormone receptor-positive and diagnosed histologically based on their distinctive discohesive growth pattern, dispersed single cells or line of cells invading the stroma. ILCs commonly show biallelic inactivation of a CDH1 (~65%), a tumor suppressor gene that is mapped to 16q22.1 and encodes for E-cadherin, a critical component of the epithelial adhesion complex. Massively parallel sequencing and pathologic studies have shown that a subset of ILCs (up to 15%) may lack CDH1 loss-of-function (LOF) mutations/deletions and retain E-cadherin expression, despite their distinctive lobular phenotype. The genetic and epigenetic underpinning of ILCs lacking CDH1 alterations has yet to be defined. Here we sought to define the mechanistic basis of the lobular phenotype in ILCs lacking CDH1 LOF genetic alterations or CDH1 gene promoter methylation and to determine the repertoire of epigenetic and genetic alterations of CDH1-wildtype ILCs. Materials and methods: Reanalysis of the CDH1 gene status in 364 primary ILCs, previously subjected to clinical massively parallel sequencing, was performed to identify all primary ILCs lacking bi-allelic CDH1 alterations. The hematoxylin and eosin stained slides of all identified ILCs lacking bi-allelic CDH1 alterations were reviewed by two pathologists and primary pure ILCs lacking bi-allelic CDH1 alterations were curated. We evaluated the presence of genetic alterations in genes playing essential roles in epithelial adhesion included in the clinical sequencing panel of up to 505 genes. The presence of CDH1 promoter methylation in 18 ILCs lacking bi-allelic CDH1 alterations with available formalin-fixed, paraffin-embedded (FFPE) material was assessed using methylation‐specific PCR (MSP) and bisulfite sequencing. Results: We identified 23/364 (6.3%) primary ILCs lacking bi-allelic CDH1 alterations, of which 65.3% and 34.7% were classic and pleomorphic lobular variants, respectively. In 18 cases with available FFPE material, our analyses revealed that 67% (12/18) of ILCs lacking bi-allelic CDH1 alterations displayed biallelic CDH1 inactivation via promoter methylation and 16q loss. Furthermore, we observed that 1/23 ILC lacking bi-allelic CDH1 alterations had bi-allelic inactivation of AXIN2. We then extended our query to all invasive breast carcinomas subjected to clinical sequencing, including the ones initially categorized as invasive breast carcinoma “type unknown”, and observed that the three additional cases with pathogenic LOF AXIN2 alterations were ILCs lacking bi-allelic CDH1 alterations. Conclusions: The lobular phenotype in ILCs can be due to CDH1 promoter methylation or genetic alterations affecting other genes related to epithelial cell adhesion, such as AXIN2 LOF mutations. Whole-genome sequencing analyses of ILCs whose molecular basis has not been identified by targeted sequencing are warranted.

Citation Format: Fatemeh Derakhshan, Higinio Dopeso, Arnaud Da Cruz Paula, Pier Selenica, Antonio Marra, Edaise M da Silva, Andrea Gazzo, Shirin Issa Bhaloo, Dara S Ross, Anne Grabenstetter, Sarat Chandarlapaty, Pedram Razavi, Hannah Y. Wen, Hong Zhang, Edi Brogi, Britta Weigelt, Fresia Pareja, Jorge S. Reis-Filho. Genetic and epigenetic basis of invasive lobular carcinomas lacking CDH1-alterations [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD14-03.