Identification Of Esr1 Splice Variants Associated With Prognosis In Estrogen Receptor Positive Breast Cancer

Background: Splice variants play a major role in carcinogenesis and disease progression. It is well known that androgen receptor splice variants are associated with resistance to prostate cancer treatment. Estrogen receptor (ER)-positive breast cancers constitute about 70% of all breast cancers and have better prognosis compared to ER-negative cancers. However, there are ER-positive breast cancers that acquire resistance to anti-estrogen therapy, and 12-55% of those tumors were shown to possess ESR1 mutations. The aim of this study was to identify common splice variants in the ESR1 gene and investigate their association with disease outcome. Methods: Whole transcriptome sequencing was performed on breast cancer specimens from 120 invasive breast cancer patients who underwent operation at Seoul National University Hospital (SNUH) and data from SNUH, GEO, and The Cancer Genome Atlas (TCGA) was used for normal breast tissue sequencing. Exon-exon junctions were identified on aligned RNA sequencing data and was used to construct exon graphs. Splice variant candidates were selected from exon graphs and were merged according to variant subtypes of samples. Subtypes were accessed differentially in relation to how frequent the junctions appear in tumor samples and common exon skipping types with frequent junctions were identified. TCGA RNA sequencing data was then used to search for the common exon skipping subtypes detected from SNUH RNA sequencing data. Results: Of the 120 tumor samples, 50 were clinically ER-positive by immunohistochemistry. Among exon paths logically possible, 125 paths were not observed in normal breast tissues. Exon 4-5 junction was the most commonly observed junction in the tumor samples. In a search for exon skipping type that results in missing ligand-binding domain of ER, three exon skipping types were identified. Exon skipping with exon 5-10 junction (type 1), exon 9-12 junction (type 2), and exon 10-12 (type 3) was seen in 4 (8%), 4 (8%), and 10 (20%) ER-positive samples, respectively. Retrospective medical chart review of the 18 patients showed recurrence in 4 (100%), 2 (50%), and 4 (40%) patients with type 1, 2, and 3 exon skipping, respectively. Evaluation of TCGA RNA sequencing data of 872 ER-positive samples suggested exon 4-5 junction as the most common junction. A search for exon skipping types in TCGA revealed 1 (0.1%), 9 (1.0%), and 454 (52.1%) samples with type 1, 2, and 3 exon skipping, respectively. However, none of the patients with type 1 or 2 had metastasis or had expired. Of the 454 patients with type 3 exon skipping, 54 patients had died, constituting 61.4% of 88 mortalities in the whole ER-positive population. Conclusion: Certain splice variants of ESR1 gene yields exon skipping subtypes commonly observed in the ER-positive breast cancer. Estrogen receptor-positive breast cancer with these exon skipping types resulting in a missing ligand-binding domain of ER may be associated with poorer disease outcome. Further investigation is warranted to validate the role of ESR1 exon skipping subtypes in the disease progression of breast cancer. Citation Format: Lee H-B, Han W, Ko S, Kim M-S, Lim S, Lee K-M, Kang YJ, Han JH, Kim Y, Yoo T-K, Moon H-G, Noh D-Y, Kim S, Han W. Identification of ESR1 splice variants associated with prognosis in estrogen receptor positive breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-04-02.