Abstract P6-03-08: Novel recurrent lncRNA fusions detected in breast cancer using RNA-Seq technology in a neoadjuvant setting
Authors: Agrawal V , Varadan V , Banerjee N , Miskimen K , Vadodkar A , Abu-Khalaf M , Sikov W , Harris L , Dimitrova N 
Published: May 2016

Background: Recent discoveries of recurrent and targetable gene fusions in breast cancer suggest the need to characterize the functional significance of such genomic aberrations within larger cohorts. We quantified fusion transcript expression in patient samples using RNASeq to identify recurrent gene fusion events in breast cancer as well as study the fusions post-brief exposure to mono-therapy.

Methods: We sequenced transcriptomes of core biopsy RNA from 130 breast tumors obtained from brief-exposure preoperative clinical trials BrUOG 211A/211B. HER2- patients were treated with brief exposure to bevacizumab (B) or nab-paclitaxel (nP) followed by treatment with B/nP/carboplatin while HER2+ patients received brief exposure to trastuzumab (T) or nP followed by T/nP/carboplatin. Paired-end sequencing on 75 baseline biopsies and 55 post-exposure biopsies using amplified total RNA yielded 55 million reads on average perlsample. Fusion transcript abundance was evaluated using 2 pipelines, TopHat-Fusion and deFuse, due to their complementary strategies in fusion detection. We eliminated gene-pseuodogene fusion pairs as likely false positives arising due to alignment artifacts. Fusions that met 1 or more of the following 3 criteria were considered high confidence:

i) Called by both deFuse and TopHat. ii) Called by deFuse with probability &gt;95% iii) Called by TopHat with &gt; 15 reads supporting the fusion.

Results: We identified high confidence gene fusions, detected by both TopHat and deFuse, in 73 of the 75 baseline biopsies with 16 fusions on average per sample. We looked for modulation of gene fusions upon brief exposure to therapy in 55 patients that had post exposure biopsy data and found that out of the 545 high confidence fusions detected across these patients, 62 (11.37%) of the fusions were found to be still present after the therapy exposure. For the recurrent fusion analyses, we considered the 75 baseline samples. We found a total of 1158 unique candidate fusions. Out of these, 116 (10%) were recurrent in more than 1 patient. After further filtering, we were able to narrow down to 9 (0.77%) fusions that were reliable since they were predicted by both the algorithms in different patients. 2 of these 9 fusions involved GAS5 as a partner gene. GAS5 have been studied to have a role in apoptosis and its down-regulation has been associated with cell proliferation, which makes it a very interesting fusion candidate.

Conclusions: We find that gene fusions in breast cancer are highly heterogeneous but are enriched with cancer-related pathway genes. This is the first study to report 2 novel gene-lincRNA fusion transcripts: MDN1-GAS5 and GABRB3-GAS5. Both these fusions are called in the baseline & post-therapy for atleast 1 patient (different patients each). GAS5 has been found as participating in a fusion in B-cell lymphoma. We are currently in the process of validating the fusion calls using qRT-PCR. The heterogeneity of detected fusions suggests that multiple mechanisms could underlie the selective advantage of tumor cells expressing fusion transcripts. The brief-exposure preoperative paradigm provides a unique opportunity to evaluate modulation of fusion transcripts that can shed light on their functional importance.

Citation Format: Agrawal V, Varadan V, Banerjee N, Miskimen K, Vadodkar A, Abu-Khalaf M, Sikov W, Harris L, Dimitrova N. Novel recurrent lncRNA fusions detected in breast cancer using RNA-Seq technology in a neoadjuvant setting. [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 P6-03-08.