BackgroundEarly-stage HR+ breast cancer patients face a prolonged risk of recurrence even after adjuvant endocrine therapy. The Breast Cancer Index (BCI) and BCIN+ prognostic models are significantly prognostic for risk of overall (0-10y) and late (5-10y) distant recurrence (DR) in N0 and N1 breast cancer, respectively. Here, the prognostic performance of BCI and BCIN+ was evaluated in a cohort of HR+ postmenopausal women from the Tamoxifen and Exemestane Adjuvant Multinational (TEAM) trial.MethodsBCI testing was performed blinded to clinical outcome with BCI/BCIN+ risk groups calculated based on pre-specified cut-points. Kaplan-Meier analysis and log-rank test were used to assess the prognostic significance of BCI/BCIN+ risk groups based on DR. Cox proportional hazard models with and without clinical covariates were used to estimate hazard ratios (HRs) and the associated 95% confidence intervals (CIs).ResultsTable: 138MOCohortNRisk groupsUnivariate HRMultivariate HRP-valueAll patientsOverall DRN0, no chemo1197Intermediate vs Low2.1 (1.4-3.4)2.2 (1.3-3.5)<0.001High vs Low4.0 (2.6-6.2)3.9 (2.5-6.3)N1, no chemo1319High vs Low2.5 (1.7-3.7)2.2 (1.5-3.3)<0.001Late DRN01285High vs Low2.1 (1.3-3.5)2.3 (1.3-3.9)0.004N11768High vs Low2.3 (1.4-3.9)2.0 (1.2-3.3)<0.001HER2-Overall DRN0, no chemo978Intermediate vs Low High vs Low2.4 (1.4 -4.0)2.5 (1.4-4.5)<0.0014.6 (2.7-7.6)5.0 (2.8-8.8)N1, no chemo1132High vs Low2.5 (1.6-3.7)2.1 (1.4-3.3)<0.001Late DRN01063High vs Low2.2 (1.2-3.9)2.5 (1.4 -4.7)0.006N11507High vs Low2.5 (1.4-4.3)2.1 (1.2-3.7)<0.001 Open table in a new tab ConclusionsBCI and BCIN+ are significantly prognostic for risk of overall DR (0-10y) and late DR (5-10y) for N0 and N1 patients, respectively. These results provide further validation of BCI clinical utility as an aid in the decision-making for adjuvant therapies for HR+ breast cancer.Legal entity responsible for the studyBiotheranostics, Inc., a Hologic Company.FundingBiotheranostics, Inc., a Hologic Company.DisclosureJ.M. Bartlett: Financial Interests, Personal, Other, Honoraria: Nanostring Technologies, Inc., Oncology Education, Biotheranostics, A Hologic Company, MedcomXchange, Communications Inc; Financial Interests, Personal, Advisory Role: Insight Genetics, Inc., BioNTech AG, Biotheranostics, A Hologic Company, Pfizer , Rna Diagnostics Inc, oncoXchange/MedcomXchange Communications Inc., Herbert Smith French Solicitors, OncoCyte Corporation; Financial Interests, Personal, Funding: Thermo Fisher Scientific , Genoptix, Agendia, NanoString Technologies, Inc., Stratifyer GmbH, Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Patents - Jan 2017: Methods and Devices for Predicting Anthracycline Treatment Efficacy, US utility – 15/325,472; EPO – 15822898.1; Canada – not yet assigned Jan 2017: Systems, Devices and Methods for Constructing and Using a Biomarker, US utility – 15/328,108; EPO – 15824751.0; Canada – not yet assigned Oct 2016: Histone gene module predicts anthracycline benefit, PCT /CA2016/000247 Dec 2016: 95-Gene Signature of Residual Risk Following Endocrine Treatment, PCT /CA2016/000304 Dec 2016: Immune Gene Signature Predicts Anthracycline Benefit, PCT /CA2016/000305 June 2020: Use of Molecular Classifiers to Diagnose, Treat and Prognose Prostate Cancer, US Provisional 63/040.692, Disclosure Name: A Molecular Classifier for Personalized Risk Stratification for Patients with Prostate Cancer, Date: 21/08/2019: Other - Patents; Financial Interests, Personal, Other, Travel, Accomodations, Expenses: Biotheranostics, A Hologic Company, NanoString Technologies, Inc., Breast Cancer Society of Canada . J. Wong: Financial Interests, Personal, Full or part-time Employment: Biotheranostics, A Hologic Company; Financial Interests, Personal, Stocks/Shares: Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Travel, Accomodations, Expenses: Biotheranostics, A Hologic Company. G. Pond: Financial Interests, Personal, Other, Honoraria: Astra-Zeneca, Takeda; Financial Interests, Personal, Advisory Role: Merck, Profound Medical; Financial Interests, Personal, Stocks/Shares: Roche Ltd. Y. Zhang: Financial Interests, Personal, Full or part-time Employment: Biotheranostics, A Hologic Company; Financial Interests, Personal, Stocks/Shares: Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Travel, Accomodations, Expenses: Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Patents: Predicting likelihood of response to combination therapy, Registered United States of America Application no: 14/724,732 Application date: 5/28/2015 Registration No: 10,253,369 Registration date: 4/9/2019; Integration of tumor characteristics with breast cancer index, pending united States of America Application no: 15/349,915 Application date: 11/11/2016; Predicting breast cancer recurrence; Pending United States of America Application no: 14/483,108 Application date: 9/10/2014; Post-treatment breast cancer prognosis, Pending United States of America Application no: 15/298,128 Application date: 10/19/2016; Neuroendocrine tumors, Pending United States of America Application no: 15/656,998 Application date: 7/21/2017: Biotheranostics, A Hologic Company. R. Salunga: Financial Interests, Personal, Full or part-time Employment: Biotheranostics, A Hologic Company; Financial Interests, Personal, Stocks/Shares: Biotheranostics, A Hologic Company; Financial Interests, Institutional, Funding: Biotheranostics, A Hologic Company. C. Markopoulos: Financial Interests, Personal, Advisory Role: Exact Sciences. D. Rea: Financial Interests, Personal, Other, Honoraria: Daiichi Sankyo, Lilly , Novartis , Pfizer , Roche; Financial Interests, Personal, Advisory Role: Genomic Health , MSD Oncology; Financial Interests, Institutional, Funding: Celgene , Roche , Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Travel, Accomodations, Expenses: Daiichi Sankyo, Eisai , Novartis , Pfizer . C.A. Schnabel: Financial Interests, Personal, Full or part-time Employment: Biotheranostics, A Hologic Company; Financial Interests, Personal, Stocks/Shares: Biotheranostics, A Hologic Company; Financial Interests, Personal, Officer: Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Patents: Predicting likelihood of response to combination therapy, Registered United States of America Application no: 14/724,732 Application date: 5/28/2015 Registration No: 10,253,369 Registration date: 4/9/2019; Integration of tumor characteristics with breast cancer index, Pending United States of America Application no: 15/349,915 Application date: 11/11/2016; Predicting breast cancer recurrence; pending united states of america application no: 14/483,108 Application date: 9/10/2014; Post-treatment breast cancer prognosis, pending united states of america application no: 15/298,128 Application date: 10/19/2016; Neuroendocrine tumors, Pending United States of America Application no: 15/656,998 Application date: 7/21/2017: Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Travel, Accomodations, Expenses: Biotheranostics, A Hologic Company. K. Treuner: Financial Interests, Personal, Full or part-time Employment: Biotheranostics, A Hologic Company; Financial Interests, Personal, Stocks/Shares: Biotheranostics, A Hologic Company; Financial Interests, Personal, Other, Travel, Accomodations, Expenses: Biotheranostics, A Hologic Company; Financial Interests, Personal, Leadership Role: Biotheranostics, A Hologic Company. J. Bayani: Financial Interests, Personal, Other, Honoraria: Thermo Fisher Scientific , NanoString Technologies; Financial Interests, Personal and Institutional, Research Grant: Thermo Fisher Scientific . All other authors have declared no conflicts of interest. BackgroundEarly-stage HR+ breast cancer patients face a prolonged risk of recurrence even after adjuvant endocrine therapy. The Breast Cancer Index (BCI) and BCIN+ prognostic models are significantly prognostic for risk of overall (0-10y) and late (5-10y) distant recurrence (DR) in N0 and N1 breast cancer, respectively. Here, the prognostic performance of BCI and BCIN+ was evaluated in a cohort of HR+ postmenopausal women from the Tamoxifen and Exemestane Adjuvant Multinational (TEAM) trial. Early-stage HR+ breast cancer patients face a prolonged risk of recurrence even after adjuvant endocrine therapy. The Breast Cancer Index (BCI) and BCIN+ prognostic models are significantly prognostic for risk of overall (0-10y) and late (5-10y) distant recurrence (DR) in N0 and N1 breast cancer, respectively. Here, the prognostic performance of BCI and BCIN+ was evaluated in a cohort of HR+ postmenopausal women from the Tamoxifen and Exemestane Adjuvant Multinational (TEAM) trial. MethodsBCI testing was performed blinded to clinical outcome with BCI/BCIN+ risk groups calculated based on pre-specified cut-points. Kaplan-Meier analysis and log-rank test were used to assess the prognostic significance of BCI/BCIN+ risk groups based on DR. Cox proportional hazard models with and without clinical covariates were used to estimate hazard ratios (HRs) and the associated 95% confidence intervals (CIs). BCI testing was performed blinded to clinical outcome with BCI/BCIN+ risk groups calculated based on pre-specified cut-points. Kaplan-Meier analysis and log-rank test were used to assess the prognostic significance of BCI/BCIN+ risk groups based on DR. Cox proportional hazard models with and without clinical covariates were used to estimate hazard ratios (HRs) and the associated 95% confidence intervals (CIs). ResultsTable: 138MOCohortNRisk groupsUnivariate HRMultivariate HRP-valueAll patientsOverall DRN0, no chemo1197Intermediate vs Low2.1 (1.4-3.4)2.2 (1.3-3.5)<0.001High vs Low4.0 (2.6-6.2)3.9 (2.5-6.3)N1, no chemo1319High vs Low2.5 (1.7-3.7)2.2 (1.5-3.3)<0.001Late DRN01285High vs Low2.1 (1.3-3.5)2.3 (1.3-3.9)0.004N11768High vs Low2.3 (1.4-3.9)2.0 (1.2-3.3)<0.001HER2-Overall DRN0, no chemo978Intermediate vs Low High vs Low2.4 (1.4 -4.0)2.5 (1.4-4.5)<0.0014.6 (2.7-7.6)5.0 (2.8-8.8)N1, no chemo1132High vs Low2.5 (1.6-3.7)2.1 (1.4-3.3)<0.001Late DRN01063High vs Low2.2 (1.2-3.9)2.5 (1.4 -4.7)0.006N11507High vs Low2.5 (1.4-4.3)2.1 (1.2-3.7)<0.001 Open table in a new tab ConclusionsBCI and BCIN+ are significantly prognostic for risk of overall DR (0-10y) and late DR (5-10y) for N0 and N1 patients, respectively. These results provide further validation of BCI clinical utility as an aid in the decision-making for adjuvant therapies for HR+ breast cancer. BCI and BCIN+ are significantly prognostic for risk of overall DR (0-10y) and late DR (5-10y) for N0 and N1 patients, respectively. These results provide further validation of BCI clinical utility as an aid in the decision-making for adjuvant therapies for HR+ breast cancer.

Abstract Background: The use of anthracycline and taxane chemotherapy has improved overall and disease-free survival in breast cancer. However these agents have significant toxicity. In addition, breast cancers can acquire or possess intrinsic chemoresistance. It is imperative to identify patients who will benefit most from adjuvant taxane treatment and those with resistant tumours who could be spared unnecessary toxicity. Methods: A panel of in vitro derived cell lines models of taxane resistance were generated by serial culture in escalating doses of either paclitaxel or docetaxel until resistance was achieved. Taxane resistant cells were characterised by 2D growth, cell cycle and apoptosis analyses. Genomic profiling using the NanoString® platform was performed to identify differentially expressed genes. The identification of kinases which target the chemoresistant models was achieved through a small molecule kinase inhibitor screen. Effects of selected target kinases on cell proliferation, cell cycle, apoptosis and protein expression were assessed. Results: Resistant cell lines exhibited an IC50 at least 40-fold higher than that of the parental cells and displayed cross-resistance to the non-establishing taxane. Cell cycle analysis revealed taxane treatment failed to induce G2/M arrest in the resistant models. A reduced apoptotic response was demonstrated. Genomic profiling identified pathways associated with the cell cycle as being significantly altered. Dinaciclib, a CDK inhibitor of CDK1, CDK2, CDK5 and CDK9, inhibited taxane resistant cell growth with IC50s comparable to the parental lines. Upon exposure to dinaciclib, cell cycle arrest at G2/M was induced and marked apoptosis demonstrated. A reduction in cyclin B1, PLK1 and pRB was observed by western blotting. Table 1:Sensitivity of taxane resistant cell lines models to paclitaxel and docetaxelCell line modelPaclitaxel (μM)Docetaxel (μM)MDA-MB-231 Parent0.004 ± 0.0030.002 ± 0.003MDA-MB-231 PACR0.184 ± 0.030.017 ± 0.02MDA-MB-231 DOCR0.414 ± 0.0470.262 ± 0.058MCF7 Parent0.004 ± 0.00050.005 ± 0.001MCF PACR0.769 ± 0.1050.07 ± 0.02 Table 2:Gene ontology enrichment analysis of biological process terms significantly over-represented in MDA-MB-231 PACR cell line modelGO TermP-valueFDRpositive regulation of transcription from RNA polymerase II promoter1.11E-162.44E-13positive regulation of cell proliferation9.99E-161.10E-12activation of cysteine-type endopeptidase activity involved in apoptotic process1.43E-106.27E-08negative regulation of apoptotic process2.13E-095.83E-07extrinsic apoptotic signaling pathway8.33E-091.62E-06cell cycle arrest8.89E-091.62E-06positive regulation of cell migration2.83E-084.42E-06 Conclusion: In this study we identified candidate resistance-associated pathways which were differentially expressed between in vitro derived taxane resistant cell line models and the sensitive parental line. The CDK inhibitor, dinaciclib, demonstrated potent activity against the taxane resistant cell line models. Clinical validation to ascertain the role of dinaciclib as a novel therapeutic in the treatment of chemorefractory breast cancer is required. Citation Format: Taylor KJ, Lyttle N, Liao L, Gourley C, Cameron DA, Bartlett JM, Spears M. Sensitivity to cell cycle inhibitors in taxane resistant breast cancer models [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-03-11.

Abstract Introduction: Male breast cancer (MBC) is a rare disease accounting for less than 1% of all breast cancers (BC) and 1% of all cancers in males. The clinical management is largely extrapolated from female BC. Several multigene assays are increasingly used to guide clinical treatment decisions in female BC, however there is little data on the utility of these tests in MBC. Methods: Here we present the gene expression results of 380 M0, ER+ve, HER2-ve MBCs enrolled in the Part 1 (retrospective joint analysis) International Male Breast Cancer Program of 1483 patients diagnosed between 1990-2010 (Cardoso et al. Annals of Oncology, 2018). Using a custom Nanostring™ panel comprised of the genes from the commercial risk tests Prosigna®, OncotypeDx® and Mammaprint®, risk scores and intrinsic subtyping data were generated to recapitulate the commercial tests as described by Bayani and Yao et al (npjBreast Cancer, 2017). Survival outcomes by risk classification were analyzed using Cox models with time-dependent covariates when the proportional hazard assumption was not met and adjusted for clinical and treatment variables. Results: Prosigna-like risk scores identified 99 (26.1%) as low-risk, 159 (41.8%) as intermediate-risk, and 122 (32.1%) as high-risk. Using the TAILORx cut-off (25) for OncotypeDx-like risk of recurrence scoring, 158 (41.6%) were identified as low-risk, while 222 (58.4%) were identified as high-risk. MammaPrint-like results identified 175 (46.1%) as low-risk and 205 (53.9%) as high-risk. Overall, patients classified as high-risk had higher grade, more nodal involvement, larger tumors, and more frequently treated with chemotherapy than low-risk patients. Survival analyses demonstrated clear clinical utility for each test, showing patients at high-risk with poor relapse-free survival (RFS) as compared to patients classified as low-risk: Prosigna-like RFS at 3-years (HR=2.20, 95% CI, 1.28-3.80); Oncotype-like RFS at 3-years (HR=1.92, 95% CI, 1.17-3.17); MammaPrint-like RFS (HR=1.51, 95% CI, 1.00-2.27); with similar findings for distant relapse-free survival (DRFS) and overall survival (OS). Across outcomes and all gene signatures, patients with concordant Low/Low risk classification had better prognosis than those with concordant High/High risk classification. PAM50 intrinsic subtyping identified 147 (38.7%) as Luminal A, 57 (15.0%) as Luminal B, 80 (21.1%) as Her2-enriched and 96 (25.3%) as Basal-like; showing overall 34.5% concordance to clinic-pathological subtyping by central pathology (95% CI, 29.7%-39.5%). Comparison between the tests in the MBC cohort and a comparable cohort of female BC from the Tamoxifen Exemestane Adjuvant Multinational (TEAM) trial processed in the same way will be presented. Conclusion: Common transcriptomic assays designed to assess residual risk, validated in female BC, provide similar information in male BC patients. Not surprisingly, disagreement between test results at the individual patient level was observed. To our knowledge, this is the largest study of MBC assayed to generate risk scores of the current commercial BC tests to demonstrate their clinical utility and their differences and similarity to female BC. This work has been funded by the Breast Cancer Research Foundation (BCRF). Citation Format: Bayani J, Poncet C, Yao CQ, Crozier C, Anouk N, Piper T, Cunningham C, Sobol M, Aebi S, Benstead K, Bogler O, Dal Lago L, Fraser J, Hilbers FH, Hedenfalk I, Korde L, Linderholm B, Martens J, Middleton L, Murray M, Kelly C, Nilsson C, Nowaczyk M, Peeters S, Peric A, Porter P, Schröder C, Rubio IT, Ruddy KJ, van Asperen C, Van Den Weyngaert D, van Deurzen C, van Leeuwen-Stok E, Vermeij J, Winer E, Boutros PC, Giordano SH, Cardoso F, Bartlett JM. Evaluation of multiple transcriptomic gene risk signatures in male breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P6-19-01.

Abstract Background:Multi-parameter tumour gene expression assays (MPAs) are widely used to estimate individual patient residual risk and to guide chemotherapy use in hormone-sensitive, HER2-negative early breast cancer. The TAILORx trial supports MPA use in a node-negative population. Evidence for MPA use in node-positive breast cancer is limited. OPTIMA (Optimal Personalised Treatment of early breast cancer usIng Multi-parameter Analysis) (ISRCTN42400492) aims to validate MPAs as predictors of chemotherapy sensitivity in a largely node-positive breast cancer population where prospective RCT (Randomised Controlled Trial) evidence is lacking. Methods: OPTIMA is a partially blinded multi-center RCT with an adaptive two-stage design. The main eligibility criteria are women and men age 40 or older with resected ER-positive, HER2-negative invasive breast cancer and up to 9 involved axillary lymph nodes. Randomisation is to standard management (chemotherapy and endocrine therapy) or to MPA-directed treatment using the Prosigna (PAM50) test. Those with a Prosigna tumour score (ROR_PT) &gt;60 receive standard management whilst those with a low score (≤60) are treated with endocrine therapy alone. Endocrine therapy for pre-menopausal women includes ovarian suppression. The co-primary outcomes are (1) Invasive Disease Free Survival (IDFS) and (2) cost-effectiveness of test-directed treatment. Secondary outcomes include IDFS in patients with low-score tumours and quality of life. An integrated qualitative recruitment study addresses challenges to consent and recruitment and will build on experience from the feasibility study that a multidisciplinary approach at sites is important for recruitment success. Tumour blocks will be banked to allow evaluation of additional MPA technologies. Recruitment of 4500 patients over 5 years will permit demonstration of 3% non-inferiority of test-directed treatment, assuming 5-year IDFS of 85% with standard management, equivalent to a HR of 1.22. Inclusion of patients from the feasibility study will increase the power to test for non-inferiority. Results: The OPTIMA main trial opened in January 2017. Overall recruitment (including the feasibility study) will reach 1000 in August 2018. Recruitment in Norway will commence in July 2018. Characteristics of the OPTIMA main participants recruited to 31st May 2018 are shown in the table. Main study patient characteristicsCharacteristic %Median age in years (range)57 (40-80) Menopause statusPre34 Post66 Male1Tumour size&lt;30mm58 &gt;=30mm42Node statuspN04 pN1mi(sn)7 pN1(sn)20 pN155 pN214Historic grade16 258 336 Conclusion: OPTIMA is one of two large scale prospective trials validating the use of test-guided chemotherapy decisions in node-positive early breast cancer. It is expected to have a global impact on breast cancer treatment. Experience from the preliminary study and close engagement with centres will aid trial success. Funding: OPTIMA is funded by the UK NIHR HTA Programme (10/34/501). Views expressed are those of the authors and not those of the HTA Programme, NIHR, NHS or the DoH. Citation Format: Stein RC, Hughes-Davies L, Makris A, Macpherson IR, Conefrey C, Rooshenas L, Pinder SE, Thomas J, Hall PS, Cameron DA, Earl HM, Naume B, Poole CJ, Rea DW, MacIntosh SA, Harmer V, Morgan A, Hulme C, McCabe C, Stallard N, Higgins H, Donovan JL, Bartlett JM, Marshall A, Dunn JA. OPTIMA: A prospective randomized trial to validate the clinical utility and cost-effectiveness of gene expression test-directed chemotherapy decisions in high clinical risk early breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr OT1-05-02.

Abstract Background: Chromosome instability (CIN) in solid tumours is associated with poor prognosis and results in numerical and structural chromosomal aberrations. Our group previously have developed the CIN signatures and have demonstrated the CIN signatures as prognostic biomarkers in breast cancer cohorts. Furthermore, our work in the BR9601 and MA.5 clinical cohorts CIN4 provided level IIB evidence that CIN4 was predictive of anthracycline sensitivity. An analysis of the DBCG 89D clinical trial was now performed to validate the role of CIN gene expression signatures as a marker of anthracycline sensitivity. Methods: RNA was extracted from patients in DBCG 89D clinical trial analysed through NanoString technology. The prognostic and predictive values of the signatures on distant relapse-free survival (DRFS) were explored using Cox proportional hazard models. Multivariate models included menopausal status, tumour size, nodal status, ER and Her2 status, histological type and grade, and treatment regimen. Results: All of the 594 samples available from the DBCG 89D we successfully analysed. CIN25 and CIN70 gene expression signatures did not associate with any of the clinicopathological characteristics tested. In addition, CIN25 and CIN70 were not prognostic or predictive of distant relapse free or breast cancer specific survival in this clinical cohort. Low CIN4 score was associated with ER negativity (p=0.02), HER2 normal expression (p<0.05). Conclusion: In this study we demonstrated that CIN4 was associated with aggressive disease. We were however in DBCG 89D unable to validate the predictive value of CIN4 concerning anthracycline sensitivity. Citation Format: Spears M, Jensen M-B, Lyttle N, Liao L, Laenkholm A-V, Ejitlertsen B, Bartlett JM. Validation of CIN4 in the DBCG 89D clinical cohort [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-11-03.

Abstract Background: Patients with high risk early breast cancer undergoing chemotherapy are frequently treated with both anthracycline and taxane-based chemotherapy exposing patients to multiple toxicities. Molecular predictors of response to specific chemotherapy agents are emerging. Abnormal duplication of the centromeric region of chromosome 17 (CEP17) and either Topoisomerase 2A (TOP2A) gene amplification or deletion, have been identified in a recent meta-analysis as potent markers of anthracycline sensitivity (Bartlett et al JCO 2015). The ROSCO study has been designed to prospectively test the utility of these markers to select anthracycline or taxane based chemotherapy in the neoadjuvant setting. Key entry criteria: Confirmed invasive breast cancer; centrally confirmed CEP17 duplication and TOP2A status; primary tumour>2cms or documented axillary node metastasis Exclusion criteria include breast cancer with good risk features i.e. Grade 1/2 ER PR rich (Q score 7/8), HER2 negative tumours. Study treatment: Patients will undergo central testing for CEP-17 and TOP2A and be randomised to 4 cycles of FEC100 (5-Flourouracil 500mg/m2 Epirubicin 100mg/m2 cyclophosphamide 600mg/m2) or 4 cycles of TC (Docetaxel 75mg/m2 Cyclophosphamide 600 mg/m2). Following chemotherapy patients will undergo surgical resection, (Institutional standard). Patients with residual invasive cancer will receive 4 cycles of the alternative chemotherapy to that received in the neoadjuvant setting. HER2 positive patients will receive concurrent trastuzumab and continue standard adjuvant trastuzumab. . Patients with biopsy proven axillary node metastases will undergo combined blue dye and radioisotope tracer guided sentinel lymph node biopsy (SLNB) and axillary lymph node clearance as a single procedure during breast surgery. Adjuvant endocrine and radiation therapy will be Institutional standard Endpoints: The primary endpoint is pathological complete response (no invasive disease in breast or axilla (pCR)). Secondary endpoints include: clinical and radiological response; rate of breast conservation; patient reported outcomes; safety, tolerability and long term outcomes. In patients with proven nodal involvement the false negative rate of a negative post treatment SLNB compared to axillary node clearance will be reported. Sample size and stratification: 1050 patients will be randomised in a 1:1 ratio stratified by nodal status, ER, HER2, and biomarker status (CEP-17 amplified or TOP2A amplified/deleted) vs normal (CEP-17 normal TOP2A normal). Analysis: The primary analysis will assess the interaction between the treatment effect and CEP17/TOP2A status to determine if a differential treatment effect exists between CEP17/TOP2A Normal and CEP17/TOP2A Abnormal patients. pCR will be analysed using a logistic regression model including co-variates for treatment, CEP17/TOP2A status and an interaction term of the two effects Sample size is based on the ability to detect an absolute improvement in pCR in the biomarker abnormal group from 21% in the TC treated group to 30% in the FEC treated group. With 90% power at 10% significance level. Contact Information: the ROSCO Trial Office ROSCO@trials.bham.ac.uk. Citation Format: Rea DW, Haywood L, Francis AM, Bowden SJ, Brookes CM, MacKenzie M, Cameron D, Stein R, Earl HM, Thomas J, Abraham J, Stanley A, Starczinski J, McGoldrick T, Treharne-Jones P, Billingham L, Bartlett JM. ROSCO: A randomised phase III, stratified CEP17/TOP2A biomarker trial of neo-adjuvant 5-flourouracil, epirubicin and cyclophosphamide vs docetaxel and cyclophosphamide chemotherapy. [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 OT3-02-02.

Abstract Background A growing number of studies are assessing the effects of drugs on breast cancers by investigating the molecular changes in tumours between the initial core biopsy taken at diagnosis and the excision specimen removed at surgery 2-3 weeks later. It is imperative to know if molecular profiles of tumours change over time without treatment and to assess whether the core biopsy itself influences the molecular profile. This study aimed to investigate changes in molecular profiles of breast cancer between the initial core biopsy and the excision specimen in patients who had no intervening treatment. Methods 83 patients with paired fresh frozen tissue specimens from an initial diagnostic core biopsy and the later surgical excision biopsy have been studied. Following RNA extraction and processing, Illumina HT-12 BeadArrays were used for gene expression profiling. Samples were analysed for significant changes in gene expression and compared with a large dataset of patients treated with neoadjuvant letrozole. Results Analysis is on-going and results from all patients will be available by December 2013. Pairwise Rank Product analysis (false discovery rate 0.01) of the first 12 patients representing all subtypes of breast cancer (6 ER+, 4 HER2+, 2 triple negative) with a mean interval of 22.9 days (range 15 - 33 days) between specimens, identified 346 down regulated genes. Functional processes represented by these genes were; ribosomes, regulation of cell death and apoptosis, and protein complex biogenesis. Up-regulated processes (236 genes) included functions relating to regulation of transcription, mRNA processing/splicing and ECM remodelling. No consistent changes in a 60 gene immune signature were seen across these 12 patients. Integration of this dataset with a large letrozole-treated dataset, demonstrated that less than 1% of genes that changed on-treatment with letrozole, changed without treatment. Conclusions This is the largest study investigating molecular profiles of the same breast cancer sampled at different times. It has shown: · No consistent changes in expression of genes associated with wound healing and immune response 15-33 days after core biopsy · Stability of the genetic profile of individual tumours despite multiple biopsies · Stability in the genes that are changed by two weeks of treatment with daily letrozole. This study provides definitive evidence of the validity of preoperative or window of opportunity molecular studies and has major implications for investigating the effects of new therapeutic drugs. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-05-05.

12 Background: Breast cancer can be classified into molecular subtypes that have distinct survival patterns. The purpose of this study was i) to evaluate the prognostic significance of breast cancer subtypes in a cohort of women taking part in the NEAT and BR9601 clinical trials comparing CMF with ECMF, and ii) to evaluate whether the subtypes were predictive of the added benefit of epirubicin in these trials. Methods: Tumor tissue microarrays were stained and scored for ER, PR, HER2, EGFR and CK5/6. These were used to classify the tumors into six intrinsic subtypes (1). We used Cox regression to compare overall survival (OS), breast cancer specific survival (BSS) and relapse free survival (RFS) in the different subgroups. We also compared the effect of ECMF with CMF by subgroup. Results: IHC data were available for 1725 cases of whom 805 were Luminal 1-basal negative, 153 were Luminal 1-basal positive, 174 were Luminal 2, 192 were HER2-like, 230 were core basal phenotype and 171 were 5-negative phenotype. Median follow-up time was 7 years. The prognostic effects of the subtypes were similar to those reported for unselected breast cancer cases irrespective of adjuvant therapy (Blows FM, et al. PLoS Med 2010;75:e1000279.). In particular, the luminal 1-basal negative tumors were associated with the best prognosis in five years after surgery and the HER2-like tumors were associated with the poorest prognosis. ECMF has previously shown to be associated with a 33% relative risk reduction for OS compared to CMF (Poole CJ et al. N Engl J Med 2006;35518:1851-62.). There was little evidence for significant heterogeneity of effect by tumor subtype for any end point (OS P= 0.40, BSS P=0.53 RFS P=0.50). However, there was an observed trend towards the largest additional benefit from ECMF being in women with tumors of the 5-negative phenotype (OS HR=0.39 95% CI 0.21-0.73) and the smallest being in Luminal 1-basal negative tumors (OS HR=0.86 95% CI 0.64-1.16). Conclusions: In a clinical trial in which all patients received chemotherapy, we confirmed that breast cancer subtypes show distinct behaviour with differences in short and long term survival. The benefit of ECMF over CMF was statistically similar in all disease subtypes.

OBJECTIVE To determine whether the density of CD4 + and CD8 + T‐lymphocytes in a transrectal ultrasonography (TRUS) biopsy of the prostate can be used to predict the progression of lower urinary tract symptoms (LUTS) in benign prostatic hyperplasia (BPH). PATIENTS AND METHODS In total, 100 patients were randomly selected from a pool of patients with histologically proven, benign TRUS biopsy specimens. There were seven full years of follow‐up available. Clinical data were recorded, including prostate volume, International Prostate Symptom Score (IPSS), prostate‐specific antigen, urine flow rate, postvoid residual urine volume and previous prostate surgery. Markers of disease progression included the subsequent development of acute urinary retention (AUR), ≥4 point rise in IPSS, prescription of medical therapy (α‐blocker or 5‐α‐reductase inhibitor) and bladder outlet surgery. Four patients’ specimens were unsuitable for analysis. Biopsy sections from 96 patients were immunohistochemically stained for the presence of CD4 + and CD8 + T‐lymphocytes and the density of infiltrate was assessed using random field sampling and point counting. RESULTS Some 29% of patients (28/96) did not have BPH at the time of biopsy. Of all patients, 41% (39/96) progressed, 10% of whom (4/39) did not have BPH at the time of biopsy. A further 10% (10/96) developed AUR, 7% (7/96) had a ≥4 point rise in IPSS, 33% (32/96) required medical therapy for BPH and 11% (11/96) required bladder outlet surgery. There was low correlation between CD4 + and CD8 + densities in paired sections. CD4 + and CD8 + densities did not provide any significant predictive function in the progression of BPH, nor was their any predictive association noted between CD4 + and CD8 + scores and the development of prostate cancer. Sub‐analysis did show that a threshold mean of ≥1.35 CD8 + cells per field predicted progression to AUR with a sensitivity of 60% (95% confidence interval, CI, 26.2–87.8), specificity of 73.3% (95% CI 62.6–82.2) but a positive predictive value of 20.6% (95% CI 8.0–39.7). CD4 + infiltrate density suggested a trend to general progression but without statistical significance. CONCLUSION The present study, despite certain trends, shows no evidence for an association between CD4 + and CD8 + T‐lymphocytes and the progression of LUTS in BPH.

5055 Background: Pre-chemotherapy identification of platinum-resistant EOC would facilitate use of alternative treatment strategies. Small studies have proposed ERCC1 as a platinum resistance marker. Here we evaluate the utility of quantitative ERCC1 mRNA and protein expression as biomarkers in a large cohort.METHODSTissue microarrays (TMAs) were constructed in triplicate from 470 advanced EOC patients treated with first-line platinum-based chemotherapy and with prospectively collected clinical and survival data. Following ERCC1 antibody validation (FL297 & SC17809 clones), immunohistochemistry (IHC) was performed. The intensity and frequency of positive staining were evaluated by both automated scoring and a human observer blinded to the case details. Quantitative analysis of protein and gene expression were also assessed using automated quantitative analysis (AQUA) of the TMAs and qRT-PCR from mRNA extracted from the same tissue specimens. Expression by each assay was correlated with survival using a minimum p-value method with subgroup analyses taking into account histology, grade, stage and chemotherapy.RESULTS457 samples were suitable for IHC/AQUA and 354 for mRNA. There was correlation between IHC results and AQUA (Spearman R value=0.33; p<0.0001), between IHC and mRNA (R=0.16; p=0.002) and between AQUA and mRNA (R=0.12, p=0.03). The median ERCC1 histoscore for patients with platinum-resistant disease (n=171) was 172 (maximum = 300; IQR 154-190), for platinum-sensitive EOC (n=286) was 177 (ns; IQR 160-194) and platinum-refractory disease subset (n=97) 169. No statistically significant difference was identified, either for PFS or OS, for any subgroup analyzed. No trend towards shorter survival in those with higher ERCC1 expression was seen using any assay.CONCLUSIONSSeveral small studies have suggested that ERCC1 expression, quantified by either IHC or qRT-PCR, can predict platinum resistance. In this, by far the largest cohort, and using quantitative methodologies, we were unable to demonstrate any association with clinical outcome. Using these assays, ERCC1 as a single gene biomarker is not useful in EOC.

TPS116 Background: Patients with HER2 +ve breast cancer suitable for neoadjuvant chemotherapy (NAC) have been shown in a series of clinical trials to have the best outcome when treated with anthracyclines (A), taxanes (T), and trastuzumab (Tz). Recent evidence confirms that adjuvant Tz is more effective when given concomitantly rather than sequentially with T (Perez SABCS 2009). Whilst there remains uncertainty as to the most efficacious A-T regimen and duration of Tz, there is widespread use in Europe of FEC-D [3 cycles of 5-FU 500 mg/m2, epirubicin 100 mg/m2 and cyclophosphamide 500 mg/m2 (FEC100) followed by 3 cycles of docetaxel 100 mg/m2 (D) q3w] following the results of PACS-01. The advent of TKI anti-HER2 agents such as L could offer superior outcomes if combined with NAC. However, a phase I study in heavily pre-treated advanced breast cancer reported difficulties in combining lapatinib (L) with D 100 mg/m2 (LoRusso JCO 2008). Methods: EORTC 10054 is designed as a two-part study to compare FEC-D wi...

Ovarian cancer is the most frequent cause of death from gynaecological cancer in the Western world. Current prognostic factors do not allow reliable prediction of response to chemotherapy and survival for individual ovarian cancer patients. Epidermal growth factor receptor (EGFR) and HER-2/neu are frequently expressed in ovarian cancer but their prognostic value remains unclear. In this study, we investigated the expression and prognostic value of EGFR, EGFR variant III (EGFRvIII), HER-2/neu and important downstream signalling components in a large series of epithelial ovarian cancer patients. Immunohistochemical staining of EGFR, pEGFR, EGFRvIII, Her-2/neu, PTEN (phosphatase and tensin homologue deleted on chromosome 10), total and phosphorylated AKT (pAKT) and phosphorylated ERK (pERK) was performed in 232 primary tumours using the tissue microarray platform and related to clinicopathological characteristics and survival. In addition, EGFRvIII expression was determined in 45 tumours by RT–PCR. Our results show that negative PTEN immunostaining was associated with stage I/II disease ( P =0.006), non-serous tumour type ( P =0.042) and in multivariate analysis with a longer progression-free survival ( P =0.015). Negative PTEN staining also predicted improved progression-free survival in patients with grade III or undifferentiated serous carcinomas ( P =0.011). Positive pAKT staining was associated with advanced-stage disease ( P =0.006). Other proteins were expressed only at low levels, and were not associated with any clinicopathological parameter or survival. None of the tumours were positive for EGFRvIII. In conclusion, our results indicate that tumours showing negative PTEN staining could represent a subgroup of ovarian carcinomas with a relatively favourable prognosis.

555 Background: There are a few large population-based series assessing the long-term outcomes of breast-conserving therapy (BCT). Methods: A consecutive series (1981–98) of 1,812 women managed by wide local excision and postoperative radiotherapy (45Gy in 20–25 fractions). Results: Mean age was 54.9 years, range 24–91. Median clinical tumor size was 20mm. Histology was no special type in 1,477 patients (78.8%), tubular in 95 patients and lobular in 121. Tumor grade was available in 1,734 patients (450 grade 1, 767 grade 2, and 517 grade 3). Of 1,725 patients with margin status recorded, there was involvement by IBC in 250 and DCIS in 50. Axillary surgery was node sample in 1,385 patients (76.4%), level III clearance in 391 patients, and level II dissection in 36. Axillary lymph nodes were histologically node negative in 1,318 patients (72.7%). Adjuvant systemic therapy was tamoxifen alone in 1,179 patients (65.1%) and other hormone therapy in 250 patients. 306 patients received chemotherapy with half these patients also receiving hormone therapy. Minimum follow-up was 7 years (median 10 years). There were 572 deaths, 329 with breast cancer present. Actuarial estimates of survival from all causes were 88.5% at 5 years and 77.2% at 10 years. Actuarial cause specific survival (CSS) was 91.2% at 5 years (95% CI 89.9%-92.5%) and 85.1% at 10 years. Significant (p<0.0001) prognostic factors for CSS were number of positive nodes, tumor grade, and pathologic tumor size. The relapse rate in the treated breast was 4.5% at 5 years, 8.5% at 10 years, 14.6% at 15 years and 19.2% at 20 years. The number of positive nodes, pathologic tumor size, and age were significant prognostic factors for breast relapse. Conclusions: Poor local control in women aged <40 questions the appropriateness of BCT in this age group. A constant local relapse rate of just under 1% per year throughout 20 years of follow-up suggests that follow-up should continue beyond 10 years. Age Number of patients 5-year ipsilateral breast relapse (%) <30 15 40.7 30-39 134 13.1 40-49 380 4.6 50-59 651 3.5 60-69 484 2.2 70- 148 5.1 No significant financial relationships to disclose.