Novel fusion genes identified from matched primary and recurrent breast cancers by RNA-sequencing

abstracts

Annals of Oncology

315P

FGFR1 and CCND1 gene amplifications are associated with breast cancer resistance to aromatase inhibitors

E. Imyanitov1, M. Kramchaninov2, T. Akhapkina1, T. Sokolova1, S. Aleksakhina1, A. Togo1, A. Iyevleva1 1 Department of Tumour Growth Biology, N.N. Petrov Institute of Oncology, St. Petersburg, Russian Federation, 2Department of Therapy, St. Petersburg City Cancer Center, St. Petersburg, Russian Federation Background: Several lines of evidence suggest the involvement of CCND1 and FGFR1 genes in determining breast cancer (BC) resistance to endocrine therapy, however these assumptions still require a validation in clinical data sets. Methods: This study included 138 tumors from patients with metastatic BC who received first-line endocrine therapy with aromatase inhibitors (AI, n ¼ 69), tamoxifen (n ¼ 65), goserelin (n ¼ 2) or a combination of goserelin and tamoxifen (n ¼ 2). DNA extracted from formalin-fixed paraffin-embedded archival specimens was tested for CCND1 and FGFR1 amplification by digital droplet PCR. Results: CCND1 and FGFR1 status was successfully determined in 134 tumors. CCND1 and FGFR1 amplification was detected in 24 (18%) and 28 (21%) informative cases, respectively; 9 carcinomas had concurrent alterations of two genes. Amplifications were more common in less differentiated tumors (G1: 1/18 (6%) vs. G2-3: 34/86 (40%); p ¼ 0.005, Fisher’s exact test). Median disease-free survival in patients receiving AI with CCND1 amplification was shorter than in cases with the normal gene status (12.3 vs. 14.9 months; p ¼ 0.014, log rank test). Objective response to aromatase inhibitors was observed in 2/13 (15%) BC with FGFR1 amplification compared to 22/46 (48%) tumors with the normal FGFR1 gene copy number (p ¼ 0.054). Noteworthy, among patients receiving AI, CCND1 and/or FGFR1 amplification occurred in 5 out of 7 (71%) women with progressive disease compared to only 4 in 23 (17%) patients with objective response to therapy (p ¼ 0.01). Meanwhile, none of 5 tumors showing resistance to tamoxifen harbored CCND1 or FGFR1 amplification. Conclusions: The presence of CCND1 and/or FGFR1 amplification is associated with worse results of AI therapy in breast cancer patients. Legal entity responsible for the study: The authors. Funding: Russian Foundation for Basic Research (grant 17-04-01281). Disclosure: All authors have declared no conflicts of interest.

Volume 30 | Supplement 5 | October 2019

316P

Clinicopathologic features of BRCA mutated breast cancer patients: Hacettepe experience

S. Aksoy1, R.F. Degirmenciler2, B.Y. Aktas1, D.C. Guven1, O.H. Aktepe1, G. Guner1, H. Taban1, N. Kertmen1, Z. Arik1, O. Dizdar1 1 Department of Medical Oncology, Hacettepe University Faculty of Medicine, Ankara, Turkey, 2Department of Internal Medicine, Hacettepe University Faculty of Medicine, Ankara, Turkey Background: Impaired DNA damage response (DDR) mechanisms and subsequent genomic instability is associated with carcinogenic process itself, but it also results in sensitivity of tumor cells to certain drugs and can be exploited to treat cancer by inducing deadly mutations or mitotic catastrophe. As a result, BRCA1/2 testing is recommended for a group individuals with breast/ovarian cancer. Methods: In this study, we retrospectively invesitigated clinicopathologic features of 303 breast cancer patients tested for BRCA 1/2 mutation in an oncology institute. NCCN recommendations are used to select patients to be tested for BRCA mutations. Results: A total of 303 patients were analysed for BRCA 1/2 mutations. Median age of the patients at diagnoses were 40 (21-65). Family history for BRCA related cancers were detected in 120 patients (39.6%). BRCA 1/2 mutations have been shown in 98 of 303 patients (32.3%). Of 98 BRCA mutated patients, 43 had pathogenic BRCA1 mutation, 39 had pathogenic BRCA2 mutation, 11 had variant of uncertain significance (VUS) BRCA2 mutation, 3 had VUS BRCA1 mutation and 2 had pathogenic BRCA1 and 2 mutations. Molecular subtypes of tumors have been shown in Table.

Table: 316P Molecular subtypes of tumors BRCA Status

HR(þ)/ HER2(-), N(%)

HR(þ)/ HER2(þ), N(%)

HR(-)/ HER2(þ), N (%)

Triple(-), N (%)

N/A,N(%)

BRCA1 BRCA2 BRCA1 VUS BRCA2 VUS BRCA1 þ 2 BRCA mut (-) TOTAL

18 (41.8%) 25 (64.1%) 2 (66.7%) 11 (100%) 0 126 (61.4%) 182 (60%)

2 (4.6%) 3 (7.7%) 0 0 1 (50%) 25 (12.1%) 31 (10.2%)

0 0 0 0 0 12 (5.8%) 12 (3.9%)

22 (51.1%) 6 (15.3%) 1 (33.3%) 0 1 (50%) 32 (15.6%) 62 (20.4%)

1 (2.3%) 5 (12.8%) 0 0 0 10 (4.8%) 16 (5.2%)

Twelve of 98 BRCA mutated patients (12.2%) were de novo metastatic while 15 patients (15.3%) faced with metastatic disease during follow-up. In the BRCA mutated group there was significantly high rate of metastatic disease has been detected comparing BRCA non-mutated group (27.5% vs %16.5% respectively, p ¼ 0.026). Metastatic BRCA mutant patients showed worse OS numerically but data is not statistically significant (P ¼ 0.894). Nine patients were diagnosed with contralateral breast cancer, and 3 patients was diagnosed with BRCA-related cancers during follow-up. Prophylactic surgery (contralateral mastectomy and/or oophorectomy) was performed in 32 of 98 patients. Conclusions: According to this retrospective study, BRCA mutant patients tend to have worse clinical and pathological features comparing age-matched controls. Legal entity responsible for the study: Sercan Aksoy. Funding: Has not received any funding. Disclosure: All authors have declared no conflicts of interest.

317P

Novel fusion genes identified from matched primary and recurrent breast cancers by RNA-sequencing

S. Choi1, J. Kim1, I.Y. Chung1, W.G. Jo1, K.W. Yun2, H.J. Park1, S.B. Lee1, H.J. Kim3, B.S. Ko3, J.W. Lee3, B.H. Son1, S.H. Ahn1 1 Department of Surgery, University of Ulsan, College of Medicine, Asan Medical Center, Seoul, Republic of Korea, 2Department of Surgery, Gangneung Asan Hospital, Gangneung, Republic of Korea, 3Breast Surgery, Asan Medical Center, Seoul, Republic of Korea Background: Breast cancers display substantial inter/intra-tumor heterogeneity. While numerous fusion genes have been identified, most are found to be subclonal, passenger alterations. To discover fusion genes which drive tumor progression and metastasis, we performed RNA-sequencing of matched primary and metastatic breast cancer samples. Methods: RNA-sequencing was performed from sixteen patients matched primaryrecurrent tumor tissue and RNA-sequencing data was successfully achieved from sixteen primary tumors and eight recurrent tumors. DeFuse program was used to identify fusion transcripts (FT). Results: Among the sixteen patients, six were hormone receptor positive, three were HER2 positive and seven were triple negative tumors. Three cases displayed locoregional recurrence only and other patients had distant metastases. Overall, 516 fusion transcripts were identified. Mean numbers of fusions in primary and recurred tumors

doi:10.1093/annonc/mdz242 | v109

Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_5/mdz242.012/5576909 by guest on 25 October 2019

(institution): Calithera Biosciences; Research grant / Funding (institution): Genentech; Research grant / Funding (institution): Medivation; Research grant / Funding (institution): Novartis; Research grant / Funding (institution): OncoSec; Research grant / Funding (institution): Pfizer Inc. J. Ettl: Honoraria (institution), Advisory / Consultancy: Lilly; Advisory / Consultancy, Travel / Accommodation / Expenses: Novartis; Honoraria (self), Advisory / Consultancy, Travel / Accommodation / Expenses: Pfizer; Honoraria (institution), Advisory / Consultancy: Roche; Advisory / Consultancy: Tesaro; Advisory / Consultancy: Eisai; Research grant / Funding (institution), Travel / Accommodation / Expenses: Celgene; Honoraria (self): TEVA; Honoraria (self): AstraZeneca. J. Balma~ na: Advisory / Consultancy, Research grant / Funding (institution), Travel / Accommodation / Expenses: AstraZenca; Advisory / Consultancy: Clovis; Advisory / Consultancy: Tesaro; Advisory / Consultancy: BMS; Research grant / Funding (institution), Travel / Accommodation / Expenses: PharMar. P.A. Fasching: Honoraria (self), Advisory / Consultancy: Roche; Honoraria (self), Advisory / Consultancy: Amgen; Honoraria (self), Advisory / Consultancy, Research grant / Funding (institution): Novartis; Honoraria (self), Advisory / Consultancy: Pfizer; Honoraria (self), Advisory / Consultancy: Celgene; Honoraria (self), Advisory / Consultancy: Puma; Honoraria (self), Advisory / Consultancy: Daiichi Sankyo; Honoraria (self), Advisory / Consultancy: Eisai; Honoraria (self), Advisory / Consultancy: Merck; Honoraria (self), Advisory / Consultancy: AstraZeneca; Honoraria (self), Advisory / Consultancy: Teva; Honoraria (self), Advisory / Consultancy: Hexal; Honoraria (institution), Speaker Bureau / Expert testimony: Myelo; Research grant / Funding (institution): BioNTech. S.A. Hurvitz: Research grant / Funding (institution): Ambryx; Research grant / Funding (institution): Amgen; Research grant / Funding (institution): Bayer; Research grant / Funding (institution): BI Pharma; Research grant / Funding (institution): Biomarin; Research grant / Funding (institution): Cascadian; Research grant / Funding (institution): Daiichi Sankyo; Research grant / Funding (institution): Dignitana; Research grant / Funding (institution): Genentech; Research grant / Funding (institution): GSK; Research grant / Funding (institution), Travel / Accommodation / Expenses: Lilly; Research grant / Funding (institution): Macrogenics; Research grant / Funding (institution): Medivation; Research grant / Funding (institution): Merrimack; Research grant / Funding (institution), Travel / Accommodation / Expenses: Novartis; Research grant / Funding (institution), Travel / Accommodation / Expenses: OBI Pharma; Research grant / Funding (institution): Pfizer; Research grant / Funding (institution): Pieris; Research grant / Funding (institution): PUMA; Research grant / Funding (institution): Roche. L.A. Albacker: Full / Part-time employment: Foundation Medicine Inc. G.M. Frampton: Full / Part-time employment: Foundation Medicine Inc. J. Chelliserry: Shareholder / Stockholder / Stock options, Full / Part-time employment: Pfizer Inc. P. Bycott: Research grant / Funding (self), Shareholder / Stockholder / Stock options, Full / Part-time employment: Pfizer Inc. U. Conte: Shareholder / Stockholder / Stock options, Full / Part-time employment: Pfizer Inc. A.M. Wardley: Honoraria (self), Advisory / Consultancy: AstraZeneca; Honoraria (self), Advisory / Consultancy: Lilly; Honoraria (self), Advisory / Consultancy: Novartis; Honoraria (self), Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Roche. M.E. Robson: Honoraria (self), Advisory / Consultancy, Research grant / Funding (institution), Travel / Accommodation / Expenses: AstraZeneca; Advisory / Consultancy: McKesson; Research grant / Funding (institution): AbbVie; Research grant / Funding (institution): BioMarin; Research grant / Funding (institution): Medivation; Research grant / Funding (institution): Myriad Genetics; Research grant / Funding (institution): Tesaro. All other authors have declared no conflicts of interest.

abstracts

318P

Association between PIK3CA mutation status and development of brain metastases in HR1/HER2- metastatic breast cancer

D.M. Fitzgerald1, A. Muzikansky2, C. Pinto2, L. Henderson3, C. Walmsley3, R. Allen4, G.B. Ferraro3, S. Isakoff5, B. Moy3, K. Oh2, H.A. Shih2, D. Dias-Santagata3, A.J. Iafrate2, A. Bardia6, P.K. Brastianos2, D. Juric7 1 Termeer Center for Targeted Therapies, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA, 2Cancer Center, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA, 3Cancer Center, Massachusetts General Hospital, Boston, MA, USA, 4Medical Oncology, Massachusetts General Hospital, Boston, USA, 5Hematology and Oncology, Massachusetts General Hospital, Boston, MA, USA, 6 Harvard Medical School, Massachusetts General Hospital, Boston, MA, USA, 7Termeer Center for Targeted Therapies, Massachusetts General Hospital, Boston, MA, USA Background: CNS metastases is a devastating complication of breast cancer, occurring in approximately 10%–15% of patients with metastatic breast cancer (MBC) and resulting in median survival of less than one year in historic cohorts. Biological factors that govern metastases to the brain, including the role of activating PIK3CA mutations, the most common actionable alterations in HRþ/HER2- MBC, are poorly understood. Methods: In this retrospective cohort study, we determined the cumulative incidence of brain metastasis in PIK3CA mutant and PIK3CA non-mutant HRþ/HER2- MBC patients treated at the Massachusetts General Hospital and genotyped during their routine clinical care with a highly sensitive multiplexed assay for real time mutation profiling of clinical samples. Results: In the overall cohort of 307 patients, 120 patients (39.1%) had PIK3CA mutant disease and 187 patients (60.9%) had PIK3CA non-mutant disease, comparable to previously published results. Median OS from the time of diagnosis of metastatic disease was 3.96 yrs (95% CI 3.40-4.87 yrs) for PIK3CA mutant patients and 4.43 yrs (95% CI 3.82-5.32 yrs) for PIK3CA non-mutant patients, p ¼ 0.6. 22.44% of patients with HRþ/HER2- disease developed brain metastases; 30.83% of PIK3CA mutant patients and 17.11% of PIK3CA non-mutant patients developed CNS metastases, p ¼ 0.0049. Median time to the development of CNS disease was 8.61 yrs for PIK3CA mutant subset and not reached (NR) for PIK3CA non-mutant subset, p ¼ 0.0086. Among patients with CNS metastases, median OS for PIK3CA mutant patients was 0.48 yrs (95% CI 0.27-0.74) and for PIK3CA non-mutant it was 1.09 yrs (95% CI 0.39-2.27), p ¼ 0.019). Conclusions: Brain metastases are common in HRþ/HER2- MBC. This incidence of brain metastases is particularly high among patients with HRþ/HER2- tumors harboring a PIK3CA mutation, where it approaches the incidence historically seen in HER2þ MBC. Early recognition of symptoms potentially related to brain metastases is important even in HRþ/HER2- subtype of breast cancer. High incidence of brain metastases in PIK3CA mutant HRþ/HER2- MBC warrants development of blood-brain barrier penetrant agents targeting the PI3K/AKT/mTOR pathway. Legal entity responsible for the study: The authors. Funding: Has not received any funding. Disclosure: S. Isakoff: Advisory / Consultancy, Research grant / Funding (institution): Myriad; Advisory / Consultancy, Research grant / Funding (institution): Puma; Advisory / Consultancy, Research grant / Funding (institution): Immunomedics; Advisory / Consultancy, Research grant / Funding (institution): Mylan; Advisory / Consultancy, Research grant / Funding (institution): Genentech; Advisory / Consultancy, Research grant / Funding (institution): Pharmamar; Advisory / Consultancy, Research grant / Funding (institution): Abbvie; Advisory / Consultancy, Research grant / Funding (institution): OncoPep; Advisory / Consultancy, Research grant / Funding (institution): Merck; Advisory / Consultancy, Research grant / Funding (institution): AstraZeneca. A. Bardia: Advisory / Consultancy, Research grant / Funding (institution): Genentech/Roche; Advisory / Consultancy, Research grant / Funding (institution): Immunomedics; Advisory / Consultancy, Research grant / Funding (institution): Novartis; Advisory / Consultancy, Research grant / Funding (institution): Pfizer; Advisory / Consultancy, Research grant / Funding (institution): Merck; Advisory / Consultancy, Research grant / Funding (institution): Radius Health; Advisory / Consultancy, Research grant / Funding (institution): Specturm; Advisory / Consultancy, Research grant / Funding (institution): Taiho Pharmaceutical; Research grant / Funding (institution): Mersana; Research grant / Funding (institution): Innocrin; Research grant / Funding (self): Biothernostics. P.K. Brastianos: Research grant / Funding (institution): Merck; Research grant / Funding (institution): Pfizer; Research grant / Funding (institution): Bristol-Myer Squibb; Advisory / Consultancy: GenentechRoche; Advisory / Consultancy: Merck; Advisory / Consultancy: Lily; Advisory / Consultancy: TESARO; Advisory / Consultancy: AngioChem. D. Juric: Advisory / Consultancy, Research grant / Funding (self), Research grant / Funding (institution): Novartis; Advisory / Consultancy, Research grant / Funding (institution): Genentech; Advisory / Consultancy, Research grant / Funding

v110 | Breast Cancer, Metastatic

(institution): Eisai; Advisory / Consultancy, Research grant / Funding (institution): EMD Serono; Advisory / Consultancy, Research grant / Funding (institution): Syros; Advisory / Consultancy: Ipsen; Research grant / Funding (institution): Takeda; Research grant / Funding (institution): Celgene; Research grant / Funding (institution): Placon Therapeutics. All other authors have declared no conflicts of interest.

319P

The role of AXL as mechanism of resistance to trastuzumab and a prognostic factor in breast cancer HER2 positive: A translational approach

A. Adam-Artigues1, E. Tormo2, F. Rojo3, J.A. Perez-Fidalgo2, S. Zazo4, P. Gonzalezon1, Alonso4, C. Hernando1, M.T. Martınez1, V. Gambardella1, J. Poveda1, S. Sim S. Moragon1, E. Alonso5, A. Rovira7, J. Albanell6, O. Burgues5, B. Bermejo2, P. Eroles2, A. Lluch2, J.M. Cejalvo1 1 Medical Oncology Department, Biomedical Research Institute INCLIVA, Hospital Clınico Vale`ncia, University of Valencia, Valencia, Spain, 2Medical Oncology Department., Biomedical Research Institute INCLIVA, Hospital Clınico Vale`ncia, University of Valencia. Center for Biomedical Network Research on Cancer (CIBERONC), Valencia, Spain, 3 Pathology Department, IIS-Fundaci on Jime´nez Dıaz, Center for Biomedical Network on Research on Cancer (CIBERONC), Madrid, Spain, 4Pathology Department, IIS-Fundaci 5 Jime´nez Dıaz, Madrid, Spain, Pathology, Hospital Clınico Valencia, University of 6 Valencia, Valencia, Spain, Medical Oncology Department, University Hospital del Mar, Center for Biomedical Network Research on Cancer (CIBERONC), Barcelona, Spain Background: Breast cancer (BC) is a heterogeneous disease. HER2þ BC represents between 15-30% of cases. Trastuzumab (T), a monoclonal antibody, has been successfully improved clinical benefits in both adjuvant and in metastatic settings. Despite this evidence, many patients experience resistance to therapy. The objective of this study is to assess AXL as a potential mechanism of resistance and its implication as a prognostic factor. Methods: We used three cell lines with acquired resistance to T. Resistant models were generated by treating parental cells (AU565, SKR3, BT474) with constant dose of T (15mg/mL) for 6 months. Cell viability was estimated by MTT assay. Proteins were assessed by Western blot (WB) and flow cytometry and genes by qRT-PCR. AXL was downregulated by siRNA and a selective AXL inhibitor (TP-0903). The prognostic value of AXL was evaluated in primary tumor in a cohort of HER2þ BC patients treated with T in adjuvant setting from Hospital Clınico Vale`ncia (n ¼ 33). Results: Acquired resistant cell lines (RCL) maintained HER2 overexpression. Cells were more proliferative and presented an increase in stem cell-like characteristics compared to sensitive parental cell lines. There was an important up-regulation of AXL (>2.5 fold-change) and epithelial-mesenchymal transition markers (VIM, CDH2, and FN1) in RCL (p < 0.05). Sensibility to T was restored by silencing AXL and with TP0903 treatment decreasing cell viability and IC50 of T (p < 0.05). AXL expression was associated with metastasis in a cohort of HER2þ BC patients (p < 0.001). There was no difference in GAS6 (a ligand of AXL). The role of AXL was also evaluated in a public data set and it was related with worse prognosis (p < 0.001). Conclusions: Our results suggest: 1) RCL were more proliferative, more mesenchymallike and stem cell-like properties; 2) AXL was a potential mechanism of secondary resistance to T; 3) Combination therapy with AXL inhibitor plus T restored sensitivity in in vitro model with AXL overexpression; 4) AXL expression was associated with relapse in HER2þ BC patients. These results showed AXL as a prognostic factor and a potential therapeutic target in HER2þ patients with resistance to T. Legal entity responsible for the study: The authors. Funding: Has not received any funding. Disclosure: A. Lluch: Advisory / Consultancy: Novartis; Advisory / Consultancy: Pfizer; Advisory / Consultancy: Roche/Genentech; Advisory / Consultancy: Eisai; Advisory / Consultancy: Celgene. All other authors have declared no conflicts of interest.

320P

Untargeted assessment of tumour fractions in plasma for monitoring and prognostication from metastatic breast cancer patients undergoing systemic treatment

M. Balic1, C. Suppan2, I. Brcic3, V. Tiran2, D.H. Mueller2, F. Posch4, P. Ulz5, E. Heitzer5, N. Dandachi2 1 Department of Internal Medicine, LKH-Univ. Klinikum Graz, Graz, Austria, 2Division of Oncology, Department of Internal Medicine, Medical University Graz, Graz, Austria, 3 Diagnostic and Research Institute of Pathology, Medical University Graz, Graz, Austria, 4 Division of Oncology, LKH-Univ.Klinikum Graz - Universit€ atsklinik fu¨r Innere Medizin, Graz, Austria, 5Institute of Human Genetics, Medical University Graz, Graz, Austria Background: The aim of this study was to assess the prognostic and predictive value of an untargeted assessment of tumor fractions in the plasma of metastatic breast cancer patients and to compare circulating tumor DNA (ctDNA) with circulating tumor cells (CTC) and conventional tumor markers. Methods: Patients and methods: 29 patients with metastatic breast cancer were included in this single-institution observational cohort study. Blood samples were obtained at first diagnosis of metastases, during several lines of treatment, and/or at every further moment of progression/development of new metastases. We assessed tumor fractions in plasma using an untargeted mFAST-SeqS method. CTCs were

Volume 30 | Supplement 5 | October 2019

Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_5/mdz242.012/5576909 by guest on 25 October 2019

were 28 and 14.6 FTs per sample. Numbers of fusions were greater in two cases with BRCA1 pathogenic germline mutations while no significant differences were observed across subtypes. Novel inter-chromosomal fusion transcript, BCL2-ESR1, CSMD1ESR1 and HPGDS-ESR1 were found in one hormone receptor positive patient’s metastasis and/or primary tumor. All fusions resulted in preservation of the DNA-binding domain and ligand-binding domain (exon4-10) of the ESR1 gene, with high ESR1 FPKM expression value. Fusions of ERBB2-, MALAT1- and CDK6- genes were found. Among the identified FTs, three cases harbored a previously reported recurrent fusion transcript EEF1DP3-FRY. Conclusions: RNA sequencing revealed numerous fusion transcripts. Among them we found novel fusions including ESR1 fusions which need further validation and functional annotation to confirm their role in tumor progression and metastasis. Legal entity responsible for the study: The authors. Funding: Asan Medical Center. Disclosure: All authors have declared no conflicts of interest.

Annals of Oncology