Abstract 927: Targeted capture next generation sequencing of fresh frozen lobular carcinoma in situ and invasive lobular cancer identifies a common repertoire of mutations
Authors: Sakr Rita A. , Scarpa Jose V. , Schizas Michael , Giri Dilip , Brot Marina De , Towers Russell , Ng Charlotte KY , Lim Raymond , Andrade Victor P. , Weigelt Britta , Reis-Filho Jorge S. , King Tari A. 
Published: May 2015

INTRODUCTION: Epidemiologic data suggest that lobular carcinoma in situ (LCIS) is a risk factor for invasive breast cancer, whereas analyses of copy number aberrations between synchronous LCIS and invasive lobular carcinoma (ILC) have provided evidence to support its role as a non-obligate precursor of ILC. Using fresh frozen (FF) tissue samples, we sought to define the repertoire of somatic mutations in LCIS and ILC, and to compare the mutational repertoire of synchronous LCIS and ILC, and synchronous foci of LCIS.

METHODS: Patients with a biopsy proven history of LCIS, with or without ILC, undergoing surgery were prospectively enrolled on an IRB approved study and FF tissue blocks were collected. Representative lesions of LCIS and ILC were microdissected and DNA was extracted using QIAamp® DNA Micro kit (Qiagen®). Matched normal germline DNA or FF normal ducts were available for all cases. At least 50 ng of DNA was used for hybridization capture using a bait library (NimbleGen®) comprising 273 genes, either recurrently mutated in breast cancer or associated with DNA repair. Targeted massively parallel sequencing was performed on a HiSeq2000 (Illumina®). Sequencing data were aligned to the reference human genome hg19 and processed by GATK. Single nucleotide variants (SNVs) were defined on the basis of three mutation callers (i.e. Mutect, Mutationseq and GATK Haplotypecaller); insertions and deletions were identified using the GATK Haplotypecaller.

RESULTS: 47 samples of LCIS and 21 samples of ILC from 30 patients (with matched normal samples) were qualified for data analysis. The constellation of somatic mutations found in LCIS and ILC was similar, with the most frequently mutated genes being PIK3CA, CDH1, HRNR and MAP3K1 found in 25% and 52%, 23% and 38%, 15% and 19% and 10% and 9% of samples, respectively. Paired analysis of synchronous LCIS and ILCs (n=21) revealed that PIK3CA and CDH1 mutations were shared in 24% and 28% of cases respectively. Overall, in 8 (38%) pairs, the LCIS and ILC had one identical mutation in common and in 7 (33%) pairs, there were two or more common identical mutations. In 5 pairs, there were no common mutations. Paired analysis of two independent foci of LCIS from 9 breasts revealed one common mutation in each of 4 (44%) pairs (CDH1, RUNX1, SPEN, ERBB2). Interestingly, the point mutation in ERBB2 (L755S) was found in two LCIS foci from a patient who underwent prophylactic mastectomy.

CONCLUSION: The mutational spectrum of LCIS is highly similar to that of ILC. PIK3CA mutations were the most common somatic mutations in both LCIS and ILC. Further, in this study, 71% of LCIS-ILC pairs shared at least one identical mutation, providing strong circumstantial evidence that LCIS is a non-obligate precursor of ILC. The finding of shared mutations among independent foci of LCIS also raises questions about the clonal origin of these lesions.

Citation Format: Rita A. Sakr, Jose V. Scarpa, Michael Schizas, Dilip Giri, Marina De Brot, Russell Towers, Charlotte KY Ng, Raymond Lim, Victor P. Andrade, Britta Weigelt, Jorge S. Reis-Filho, Tari A. King. Targeted capture next generation sequencing of fresh frozen lobular carcinoma in situ and invasive lobular cancer identifies a common repertoire of mutations. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 927. doi:10.1158/1538-7445.AM2014-927