Prosigna Test Research Articles

Prosigna Assay for Treatment Decisions in Early Breast Cancer: A Decision Impact Study.

Introduction: Therapeutic decisions in early breast cancer are based on clinico-pathological features which are subject to intra- and inter-observer variability. This single-center decision impact study aimed to evaluate the effects of the Prosigna assay on physicians' adjuvant treatment choices. Methods: Between 09/2017 and 02/2018, formalin-fixed tumor samples from 52 newly diagnosed, postmenopausal, hormone receptor-positive, HER2-negative breast cancer (T1-T2; pN0-N1a) patients were analyzed. Pre-test clinical judgements and Prosigna test results were compared. Results: The mean age was 59 (42-77). Invasive ductal carcinoma (79.2%), grade 2 (52.8%) and T1c-N0 tumors (43.4%) were represented. There was 40.4% discordance between the pre- and post-test risk of recurrences. No significant change was observed in the clinical intermediate risk category, while there was a net reclassification of low-risk patients into a high Prosigna recurrence risk group. In addition, clinically determined intrinsic subtypes were 34.6% discordant with the Prosigna results, which is largely driven by the reclassification of the luminal A tumors into the Prosigna-assessed luminal B group. Before the Prosigna test, endocrine treatment was the primary choice in 20 patients (39.2%), and chemotherapy was recommended to 31 patients (60.8%). Overall, the Prosigna assay led to a change in treatment choice for one patient. Conclusions: Although conventional risk assessment methods are relatively inexpensive with shorter turnaround times, their accuracy and value for risk reduction are suboptimal. According to our results, the Prosigna assay was found to be a relevant tool for the clinical decision making process. Long-term follow-up of these patients will elucidate the potential benefits of using multigene molecular tests as biomarkers for treatment.

Retrospective observational study of patients with luminal breast cancer in early stages: 10-year follow-up—Which luminal breast cancer relapse?

e12586 Background: Luminal BC is a heterogeneous group of breast cancer (1), with molecular signatures of better prognoses and others of worse prognoses being found. Prosigna test provide the molecular BC subtype, an individualized prognostic score called “ROR score”. Our main objective is to analyze in 10 years which Luminal early BC patients have local and distant relapses. See if there is a correlation between ROR and relapse. As second objective is select this group of patients who relapse and analyze the presence of CAF S1 for a future study. Methods: We performed a retrospective observational study from the two main hospitals in Málaga (Spain) We included 415 women 43-79 years with EBC defined as T1 or T2 primary breast cancer <30 mm in the largest dimension) node negative. Molecular BC subtype analyzed by IHC and genomic platform Prosigna) The patients were treated with breast-conserving surgery with clear excision margins hypofractionated radiotherapy and adjuvant endocrine therapy. We followed them from 2014 to the present. Results: 415 patients. We made genetic platform Prosigna to all of them luminal A 55%, Luminal B 37%, Basal like 4.8% and Her2 3.6%. The median follow-up was 9.1 years. The cumulative incidence of recurrence in Luminal BC was 2.29% (44.4% locoregional 55.55% distant recurrence). In 55.55% of the patients that relapsed had a Prosigna test with high ROR score, in 11.11% of the patients that relapsed had a Prosigna test with an intermediate ROR score and in 33.33 of the patients that relapsed had a Prosigna test with a low ROR score. Conclusions: In our study the cumulative incidence of recurrence in Luminal BC was 2.29 % and the ROR score provided a good estimate of the risk of relapse. Currently we use genomic platforms as Prosigna to estimate the ROR and adapt treatments to the risk. However, until now there is no biological or translational study focusing on T1N0 luminal BC. Subsequently, there exists a real need for new strategies to determine the long-term prognosis of these patients. We selected a group of patients to analyze the presence of CAF S1 fibroblast for a future study. Distant recurrence in T1N0 Luminal BC is strongly associated with the presence of CAF-S1 fibroblast The determination of CAF-S1 could more accurately predict recurrence in this subgroup of patients and could be a target for future treatments.

Use of the Gene Expression Test Prosigna® in Premenopausal Patients with HR+, HER2− Early Breast Cancer: Correlation of the Results with the Proliferation Marker Ki-67

Introduction: In hormone receptor-positive (ER+/PR+) and human epidermal growth factor receptor 2-negative (HER2−) early-stage breast cancer (EBC), gene expression tests such as the Prosigna are increasingly used since classic clinicopathological parameters and the proliferation factor Ki-67 often do not allow a definite therapy decision regarding an adjuvant chemotherapy. While the Prosigna test has been validated for postmenopausal patients, few data are available regarding its use in premenopausal patients. The present study compared the Prosigna test with the Ki-67 index in premenopausal patients. Materials and Methods: Premenopausal patients with HR+ HER2−, pN0-1, G1-2 EBC were retrospectively enrolled (n = 55). The Prosigna assay was performed in formalin-fixed paraffin-embedded tumor samples of surgical resection specimens. Ki-67 was reassessed in original diagnostic core needle biopsy specimens and defined as low, intermediate, or high with the threshold of <10%, 10–24%, ≥25%. Results: According to Ki-67, patients were in the low (LR)-, intermediate (IR)-, and high-risk (HR) groups in 40%, 36%, and 24% of the cases. The Prosigna gene signature assay assessed the risk of recurrence as LR for 45% of the patients, IR for 35%, and HR for 20%. The most frequent intrinsic subtypes were luminal A in 73% and luminal B in 24% of the patients. A moderate correlation was found between Prosigna and Ki-67 scores with a Pearson correlation coefficient of 0.51. In the overall cohort, 47% of the Ki-67-based therapy decision would correspond to those based on the Prosigna score. After exclusion of IR patients, matching of low/low or high/high results was observed in 57% of the cases. Conclusion: According to the present study, there is only limited concordance regarding the risk group stratification between Ki-67 and Prosigna-based risk assessment. The relevance and frequency of premenopausal breast cancer emphasizes the need for further evaluation of gene expression analyses in this setting and the correlation with classic clinicopathological parameters regarding therapy decision-making.

Compliance to genomic test recommendations to guide adjuvant chemotherapy decision-making in the case of hormone receptor-positive, human epidermal growth factor receptor 2-negative breast cancer, in real-life settings.

Genomic tests are a useful tool for adjuvant chemotherapy decision-making in the case of hormone receptor-positive (HR+), and human epidermal growth factor receptor 2-negative (HER2-) breast cancer with intermediate prognostic factors. Real-life data on the use of tests can help identify the target population for testing. French multicentric study (8 centers) including patients, all candidates for adjuvant chemotherapy for HR-positive, HER2-negative early breast cancer. We describe the percentage of tests performed outside recommendations, according to the year of testing. We calculated a ratio defined as the number of tests required to avoid chemotherapy for one patient, and according to patient and cancer characteristics. We then performed a cost-saving analysis using medical cost data over a period of 1 year from diagnosis, calculated from a previous study. Finally, we calculated the threshold of the ratio (number of tests required to avoid chemotherapy for one patient) below which the use of genomic tests was cost-saving. A total of 2331 patients underwent a Prosigna test. The ratio (performed test/avoided chemotherapy) was 2.8 [95% CI: 2.7-2.9] in the whole population. In the group following recommendations for test indication, the ratio was 2.3 [95% CI: 2.2-2.4]. In the case of non-abidance by recommendations, the ratio was 3 [95% CI: 2.8-3.2]. Chemotherapy was avoided in 841 patients (36%) following the results of the Prosigna test. The direct medical costs saved over 1 year of care were 3,878,798€ and 1,718,472€ in the group of patients following test recommendations. We calculated that the ratio (performed test/avoided chemotherapy) needed to be under 6.9 for testing to prove cost-saving. The use of genomic testing proved cost-saving in this large multicentric real-life analysis, even in certain cases when the test was performed outside recommendations.

103P Impact of Prosigna test on treatment decision in lymph node-negative early breast cancer: A prospective multicenter study (EMIT1)

103P Impact of Prosigna test on treatment decision in lymph node-negative early breast cancer: A prospective multicenter study (EMIT1)

Abstract OT3-32-01: 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 Background: Multi-parameter tumor gene expression assays (MPAs) are used to estimate individual patient risk and 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 postmenopausal node-positive breast cancer has been provided by the RxPONDER trial interim analysis but this relies on the absence of superiority in an analysis where &amp;gt;50% of events were unrelated to breast cancer. There is much uncertainty about MPA use for premenopausal patients. OPTIMA (Optimal Personalised Treatment of early breast cancer usIng Multi-parameter Analysis) (ISRCTN42400492) is a prospective international randomized controlled trial designed to validate MPAs as predictors of chemotherapy sensitivity in a largely node-positive breast cancer population. Methods: OPTIMA is a partially blinded study with an adaptive two-stage design. The trial recruits women and men age 40 or older with resected ER-positive, HER2-negative invasive breast cancer and up to 9 involved axillary lymph nodes. Randomization is to standard management (chemotherapy and endocrine therapy) or to MPA-directed treatment using the Prosigna (PAM50) test. Those with a Prosigna tumor Score (ROR_PT) &amp;gt;60 receive standard management whilst those with a low score (≤60) tumor are treated with endocrine therapy alone. Endocrine therapy for pre-menopausal women includes ovarian suppression for all participants unless they experience a chemotherapy-induced menopause. Adjuvant abemaciclib is permitted. The trial will be analyzed for (1) non-inferiority of recurrence according to randomization and (2) cost-effectiveness. The key secondary outcome is non-inferiority of recurrence for patients with low ROR_PT score tumors. The efficacy analyses will be performed Per Protocol using Invasive Breast Cancer Free Survival (IBCFS) as the primary outcome measure to limit the risk of a false non-inferiority conclusion. Recruitment of 4500 patients over 8 years will permit demonstration of up to 3% non-inferiority of test-directed treatment with at least 83% power, assuming 5-year IBCFS is 87% with standard management. An integrated qualitative recruitment study addresses challenges to consent and recruitment, building on experience from the feasibility study which found that a multidisciplinary approach is important for recruitment success. OPTIMA is strongly supported by a patient group which has helped design all patient documents and which is represented on the TMG. Results: The OPTIMA main trial opened in January 2017 and has continued to recruit throughout the COVID-19 pandemic. Overall recruitment as of 1 July 2022 was 2814 (2593 from UK, 221 from Norway). Patient characteristics are well balanced between the trial arms. Currently 95% of randomized participants are eligible for inclusion in the PP analysis. 66% of the MPA-directed arm participants have been allocated to endocrine therapy only. The test failure rate is &amp;lt; 1%. Conclusion: OPTIMA will provide robust unbiased evidence on test-directed chemotherapy safety for both postmenopausal and premenopausal women with 1-3 involved nodes as well as for patients with 4-9 involved nodes and for patients treated with abemaciclib. Funding: OPTIMA is funded by the UK NIHR HTA Programme (10/34/501) and in Norway by KLINBEFORSK and the Norwegian Cancer Society. Views expressed are those of the authors and not those of the HTA Programme, NIHR, NHS or the Department of Health. Trial Inquiries: OPTIMA@warwick.ac.uk Patient characteristics Citation Format: Robert Stein, Andreas Makris, Iain Macpherson, Luke Hughes-Davies, Andrea Marshall, Georgina Dotchin, David A. Cameron, Belinda E. Kiely, Caroline Wilson, Anne Armstrong, Helena M. Earl, Christopher J. Poole, Janice Tsang, Bjørn Naume, Daniel Rea, Hege Ohnstad, Peter S. Hall, Stuart A. McIntosh, Bethany Shinkins, Christopher McCabe, Adrienne Morgan, John MS Bartlett, Janet A. Dunn. 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 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr OT3-32-01.

The use of clinical impact of the breast cancer intrinsic subtype-Prosigna assay for adjuvant treatment decision in early breast cancer with hormone receptor positive and HER-2 negative Middle East women.

e12527 Background: Clinico-pathological evaluations of intrinsic subtypes by immunohistochemistry (IHC) are routinely used for treatment decision in patients with early-stage breast cancer (EBC). The Prosigna (PAM50) gene signature, is validated test measures the expression levels of 50 genes from formalin-fixed paraffin-embedded (FFPE) tumor samples. It permits intrinsic subtype classification of tumor and prognostic test of risk of recurrence (ROR) score. The objectives of this study were to evaluate the impact of Prosigna results on type of adjuvant treatment in EBC, to overcome disparities by comparing the results on different ethnicity groups in Israel and to asses physicians treatment decision before and after Prosigna results. Methods: This prospective, observational study was carried out in 7 Medical Centers at Northern Israel. Postmenopausal women with hormone-receptor–positive, human epidermal growth factor receptor 2 (HER2)–negative invasive EBC were enrolled. Physicians completed treatment confidence questionnaires prior to and after receiving Prosigna test results. Only EBC : T 1-2, N0 and N1 were included. Results: Between May 2018 and April 2021, 238 women were enrolled, 204 patients are eligible for this analysis. Ashkenazi Jews were 38%, Sephardi Jews 32% and 30% were Palestinian Arab women. Mean age was 65.28 years. T1 was diagnosed in 82.3%, T2 in 17.3% and N1 in 22.4% of pts. At time of diagnosis tumor subtypes were categorized based on IHC as Luminal A in 50%, 54.4%, 59.5% and Luminal B in 50%, 44.6% and 40.5% of Sephardi, Ashkenazi and Arab patients retrospectively. Intrinsic tumor subtypes detected by the Prosigna test were Luminal A in 63.8%, 71%, 74.3% and Luminal B in 34.5% (+1.7% basal like), 29% and 25.7% respectively. No significant different between these groups. Overall 50.5% of pts were low risk, 26% intermediate risk and 23.5% high risk due to Prosigna results. Following Prosigna results 23.4% of the physicians changed their treatment recommendation. Conclusions: To our knowledge this is the first study using Prosigna test in Middle East women with early breast cancer. The use of Prosigna results led to a 23.4% change in adjuvant treatment decision. The results of this prospective study is similar to the data published in other studies in the literature. The intrinsic tumor subtypes and risk of recurrence detected by Prosigna adds to the physicians' confidence in the choice of treatment type. Prosigna test was additive for understanding the biology and for adjuvant treatment choice in both Jewish and Palestinian women in Israel.

Sample Preparation Approach Influences PAM50 Risk of Recurrence Score in Early Breast Cancer.

Simple SummaryThe PAM50 risk of recurrence (ROR) score is predictive of the risk of distant recurrence and the benefits of adjuvant therapy in early breast cancer. The Prosigna assay utilizes RNA from tumor cells selected via macrodissection of formalin-fixed, paraffin-embedded (FFPE) tissue sections to measure the activity of the PAM50 genes. An alternative and widely used extraction method is RNA purification from fresh-frozen (FF) bulk tissue without enriching the tumor cellularity. However, the impact of the RNA preparation approach on ROR scores and subsequent treatment selection has not been systematically evaluated. We compared the different approaches and found high correlation between risk of recurrence scores estimated from macrodissected FFPE tissue and scores estimated from bulk FF tumor tissue. However, important discrepancies were revealed for luminal tumors, which may have consequences for treatment recommendations for these patients.The PAM50 gene expression subtypes and the associated risk of recurrence (ROR) score are used to predict the risk of recurrence and the benefits of adjuvant therapy in early-stage breast cancer. The Prosigna assay includes the PAM50 subtypes along with their clinicopathological features, and is approved for treatment recommendations for adjuvant hormonal therapy and chemotherapy in hormone-receptor-positive early breast cancer. The Prosigna test utilizes RNA extracted from macrodissected tumor cells obtained from formalin-fixed, paraffin-embedded (FFPE) tissue sections. However, RNA extracted from fresh-frozen (FF) bulk tissue without macrodissection is widely used for research purposes, and yields high-quality RNA for downstream analyses. To investigate the impact of the sample preparation approach on ROR scores, we analyzed 94 breast carcinomas included in an observational study that had available gene expression data from macrodissected FFPE tissue and FF bulk tumor tissue, along with the clinically approved Prosigna scores for the node-negative, hormone-receptor-positive, HER2-negative cases (n = 54). ROR scores were calculated in R; the resulting two sets of scores from FFPE and FF samples were compared, and treatment recommendations were evaluated. Overall, ROR scores calculated based on the macrodissected FFPE tissue were consistent with the Prosigna scores. However, analyses from bulk tissue yielded a higher proportion of cases classified as normal-like; these were samples with relatively low tumor cellularity, leading to lower ROR scores. When comparing ROR scores (low, intermediate, and high), discordant cases between the two preparation approaches were revealed among the luminal tumors; the recommended treatment would have changed in a minority of cases.

175P Prosigna test for early breast cancer patients in real-life

Genomic tests are used as adjuvant chemotherapy decision support tool in HR + HER2- breast cancer (BC) patients. In France, the Health Authorities have recently advised indications for tests in case of intermediate risk of recurrence: pT1c-pT2 pN0-pN1mic grade 2 BC (the HA group). However, in routine practice, some tests were performed before and outside these indications. The aim of this study was to identify a population that benefits from performing a genomic test in real world setting.

Prosigna test in breast cancer: real-life experience.

Genomic tests can guide the decision to administer adjuvant chemotherapy in women with hormone receptor (HR)-positive, Human Epidermal growth Factor 2 (HER2)-negative breast cancer (BC) at intermediate risk of recurrence. We assessed the decision-making and economic impact of the Prosigna test in a real-life setting. Retrospective cohort study of HR + , HER2- BC patients managed from 2016 to 2020, potential candidates for adjuvant chemotherapy, at intermediate risk of recurrence, in whom a Prosigna test was performed according to contemporary guidelines. The additional cost of chemotherapy over one year in terms of direct medical and non-medical costs was estimated in this study to be €9,737 (derived from a previous study, NCT02813317). The cost of the Prosigna test, as defined by the reimbursement system, was €1,849. Among the 809 patients included in this study, 2.3 Prosigna tests had to be performed to avoid adjuvant chemotherapy for one patient. The number of tests that had to be performed to avoid chemotherapy for one patient was higher for patients with grade 3 tumors and pN1mic axillary node involvement and lower for grade 1 tumors or in the absence of axillary node involvement (pN0), but did not vary according to the 10-year overall survival gain predicted by the Predict online test. The cost saving related to withholding of adjuvant chemotherapy for one patient on the basis of the Prosigna test results was €5,485. We present one of the largest cohorts of HR + , HER2- BC patients at intermediate risk of recurrence, in whom a Prosigna test was used to guide the adjuvant therapy decision in a real-life setting, resulting in a 44% decrease in the indication for chemotherapy.

Abstract OT3-17-01: OPTIMA: A prospective randomized trial to validate the clinical utility and cost-effectiveness of gene expression test-directed chemotherapy decisions in mostly node-positive early breast cancer

Abstract Background: Multi-parameter tumour gene expression assays (MPAs) are widely used to estimate individual patient 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 in node-positive breast cancer is limited. OPTIMA (Optimal Personalised Treatment of early breast cancer usIng Multi-parameter Analysis) (ISRCTN42400492) is a prospective international randomised controlled trial (RCT) designed to validate MPA’s as predictors of chemotherapy sensitivity in a largely node-positive breast cancer population. Methods: OPTIMA is a partially blinded study 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) &amp;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. Prosigna tests are currently performed only for participants randomised to MPA-directed treatment. More than 1 tumour may be tested if participants have multi-focal tumours with discordant features and/or are considered clinically significant. The co-primary outcomes are: (1) Invasive Disease Free Survival (IDFS) and (2) cost-effectiveness. Secondary outcomes include IDFS in patients with low-score tumours and quality of life. 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. An integrated qualitative recruitment study addresses challenges to consent and recruitment, building on experience from the feasibility study which found that a multidisciplinary approach is important for recruitment success. Results: The OPTIMA main trial opened in January 2017. Overall recruitment as of 1 July 2019 was 1123 (1100 from UK, 13 from Norway); 91% had axillary node macro-metastases. Median time from consent to treatment allocation was 12 days (interquartile range 10-14 days). The withdrawal rate from trial treatment is 3%; 50% of these continue with follow up. Prosigna tests have been performed on 608 tumours for 549 participants; 59% were luminal A, 38% were luminal B and 3% non-luminal (6 patients with non-luminal tumours [1% overall] were ineligible on receptor retesting). Of the 53 (10%) participants with &amp;gt;1 tumour tested, 3 (6%) had discordant scores only, 7 (13%) had discordant subtypes only and 8 (15%) had both discordant scores and subtypes. Two thirds of the MPA-directed arm participants have been allocated to endocrine therapy only. The test failure rate is &amp;lt;1%. 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. 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 Department of Health. Trial Inquiries: OPTIMA@warwick.ac.uk Citation Format: Robert C Stein, Andrea Marshall, Andreas Makris, Luke Hughes-Davies, Iain R MacPherson, Carmel Conefrey, Leila Rooshenas, Sarah E Pinder, Abeer M Shaaban, Bjørn Naume, David A Cameron, Daniel W Rea, Helena M Earl, Christopher J Poole, Peter S Hall, Georgina Dotchin, Stuart A McIntosh, Victoria Harmer, Adrienne Morgan, Bethany Shinkins, Nigel Stallard, Christopher McCabe, Jenny L Donovan, John MS Bartlett, Janet A Dunn. OPTIMA: A prospective randomized trial to validate the clinical utility and cost-effectiveness of gene expression test-directed chemotherapy decisions in mostly node-positive early breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr OT3-17-01.

Abstract P3-11-10: Prosigna prognostic signature in pN1mi, estrogen receptor-positive breast cancer:the pN categorization impacts the BC risk stratification

Abstract Background Several validated molecular subtyping tests for breast cancer based on gene expression profiling are now routinely used in clinical practice.The Prosigna® breast cancer (BC) prognostic gene signature assay identifies a gene-expression profile that permits the classification of tumors into subtypes and gives a score for the risk of recurrence (ROR) at 10 years. The assay uses the 50-gene expression profile, weighted together with clinical variables, to generate a risk category and numerical score. The score is reported on a 0-100 scale, for hormone receptor positive BC. Prosigna test results classified tumors according to intrinsic subtypes (Lum A, Lum B, HER2-enriched, basal-like) and ROR risk groups (low risk, 0-40; intermediate risk, 41-60; and high risk, 61-100). For node-positive patients, the weight given to the node status is similar when considering pN1mi ([0.2-2mm]) or pN1 (&amp;gt;2mm, 1-3 positive lymph nodes). Our aim was to characterize the Prosigna test classification of the pN1mi subgroup in the French national registry for molecular signatures in ER+ BC. Methods Since 2018, four French laboratories using Prosigna test in clinical practice retrospectively implement a national registry for molecular prognostic signatures in ER+ BC. We analyzed the pN1mi subgroup with regards to their clinico-pathological characteristics, Prosigna test results and ROR risk score. Using the definition formula of the prosigna algorithm, we could calculate a hypothetical score if the pN1mi has been considered as pN0 and redefine risk categories for pN1mi breast cancers. Results The database included 886 tests performed in routine practice, including 91 (10.3%) pN1mi. The pN1mi BC were ER+ HER2- (88/91, 96.7%), invasive carcinoma of no special type in 81/91 (89%), with a median tumor size of 23 mm (range 12-60). Among these, the median ROR score was 44, and 51/91 tumors (56%) were classified as LumA tumors, 34/91 (41%) as LumB, two asHER2-enriched and one as basal-like. Interestingly, the probability of distant recurrence was 26% if pN1mi BC were considered as N+ but lowered to 10% if considered as pN0. Among the different molecular subtypes, the change from pN1 top N0 has different weight: in Lum A tumors, the switch in the probability of distant recurrences was 13 to 6% according to pN categorization of the pN1mi BC, and the median ROR score moved from 27 to 6 with 15/51(29%) of Lum A tumors considered as high risk if pN1 but only 1/51 if pN0; 31/51 (61%) versus 14/51 (27.5%) were in the intermediate risk group, and only 5/51 (10%) versus 34 (67%) in the low risk group. For LumB tumors, the switch in the probability of distant recurrences was 29% to 17% according to pN categorization and and the median ROR score moved from 60 to 15 with 36/37(97.3%) of Lum B tumors considered as high risk if pN1but only 13/37(35%) ifp N0; 1/37 versus 20/37 (54%) were in the intermediate risk group, and none of the 37 LumB versus 1 in the low risk group. Conclusion With regards to Prosigna test in pN1mi BC - and the persistent debate as to whether they should bear the same weight as pN1 pN0 tumors - categorization in ROR risk group is likely to be impacted by the node factor weight, and more importantly among Lum B tumors. Citation Format: Charafe-Jauffret E, Duprez-Paumier R, Berghian A, Penault-Llorca F, Dauplat MM, Boucraut A, Cohen M, Houvenaeghel G, Maroc C, Pradines A, Cayre A, Etancelin P, Lacroix-triki M. Prosigna prognostic signature in pN1mi, estrogen receptor-positive breast cancer:the pN categorization impacts the BC risk stratification [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 P3-11-10.

Abstract P3-08-23: Prosigna assay for treatment decisions in early breast cancer: A single center, decision impact study

Abstract Background: Therapeutic decisions in early breast cancer (EBC) are based on clinical and pathological features, which are subject to intra- and inter-observer variability. Hence, in the era of precision medicine, there is growing need for predictive biomarkers. The Prosigna assay utilizes Prediction Analysis of Microarray, a test based on the analysis of 50 intrinsic subtype-linked gene clusters. This single center decision impact study aimed to evaluate the effect of Prosigna test results on physicians' adjuvant treatment choices. Methods: Between September 2017 and February 2018, FFPE tumor samples from 53 newly diagnosed, postmenopausal, hormone receptor-positive, HER2-negative EBC (T1-T2; pN0-N1a) patients were analyzed. Pre-test clinical judgments and Prosigna test results were compared. Results: Mean age was 59 (42-77). Invasive ductal carcinoma (79.2%), grade 2 (52.8%) and T1c-N0 tumors (43.4%) represented the majority. Before the Prosigna test, 65.4% of the patients were classified as luminal A and 34.6% as luminal B. Of the pre-test risk groups, 40.4% were low-risk, 40.4% were intermediate risk and 19.2% were high risk. Prosigna assay grouped 50% of patients as luminal A, 44.2% as luminal B, 3.8% as basal type and 1.9% as HER2-expressing. Post-test ROR score-based groups were distributed as 25% low-risk, 40.4% intermediate risk and 34.6% high risk. There was a statistically significant correlation between clinically defined and molecularly assessed intrinsic BC subtypes (kappa:0.334, p=0.007). Similarly, pre-test and post-test recurrence risk groups were correlated (kappa:0.397, p=0.001). Before the Prosigna test, endocrine treatment was physicians' primary choice in 20 patients (39.2%), chemotherapy was recommended to 31 patients (60.8%). Overall, the Prosigna assay led to a change in choice of treatment for one patient (2%). There was 40.4% discordance between pre- and post-test recurrence risk groups. In addition, intrinsic subtypes were 34.6% discordant, which is largely driven by the reclassification of pre-test luminal A tumors into Prosigna luminal B group. Table 1.Impact of Prosigna Results on Final Treatment Decision Prosigna low risk N=13 (%24.5)Prosigna intermediate risk N=21 (%39.6)Prosigna high risk N=18 (%34)Total N=51 (%100)Treatment choice before Prosigna CT+HT0121729 (56.9)HT only127120 (39.2)CT offered, not accepted by the patient0202 (3.9)Treatment choice after Prosigna CT+HT0121830 (58.8)HT only127019 (37.3)CT offered, not accepted by the patient0202 (3.9)Change in treatment choice HT to CT001100 (0)CT to HT0000 (0)CT: Chemotherapy HT: Hormonotherapy Conclusions: Although conventional risk assessment methods are relatively inexpensive with shorter turnaround times, their accuracy for risk assessment and value for risk reduction are suboptimal. According to our results, Prosigna assay was found as a relevant tool for clinical decision-making process. In cases where there is a discrepancy between the clinical assessment results and the Prosigna assay, tumor boards may guide treatment recommendations. Long term follow-up of these patients will elucidate the potential benefits of using multigene molecular tests as biomarkers for EBC treatment. Citation Format: Esin E, Oksuzoglu BO, Markoc F, Bilgetekin I, Yildiz F, Guntekin S, Yukruk F, Atalay R. Prosigna assay for treatment decisions in early breast cancer: A single center, decision impact study [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 P3-08-23.

Abstract PD3-02: Incidence of late relapse in HER2-positive (HER2+) breast cancer patients receiving adjuvant trastuzumab: Combined analysis of NCCTG (Alliance) N9831 and NSABP (NRG) B31

Abstract Background: Recent trials showed potential benefit of extended adjuvant endocrine therapy and relatively high risk of relapse after 5 years (yr) in hormone receptor-positive (HR+) HER2- breast cancer. While risk of late relapse in HR+ HER2- is fairly well defined, the risk of late relapse in HER2+ remains largely unknown. Method: 4547 HER2+ patients treated with adjuvant chemotherapy alone or combined with trastuzumab (TH) were included (3132 from North Central Cancer Treatment Group [NCCTG, now Alliance] N9831; 1415 from National Surgical Adjuvant Breast and Bowel Project [NSABP, now NRG] B-31). Intrinsic subtypes were assessed by Prosigna test. Kaplan-Meier method and Cox proportional hazards model were used for analysis. Results: Median follow-up was 10.4 yr in N9831 and 7.0 yr in NSABP-B31. 54.5% of pts had HR+ disease. Pts were classified as HER2 enriched (77.4%), Luminal B (10.1%), Luminal A (7.7%), and Basal (4.8%). In multivariate Cox regression analysis in both treatment groups, HR+ was significantly associated with improved recurrence-free survival (RFS) during the first 5 yr (HR 0.65, 95% CI 0.56-0.77, p&amp;lt;0.001) but there was no significant difference during yr 5-10 (HR 1.32, 95% CI 0.93-1.88, p=0.12). In HR+HER2+ pts treated with TH, cumulative hazard for relapse or death was lower in the first 5 yr (10.96%, 95%CI 8.78-13.08) compared to HR-HER2+ (17.48%, 95%CI 14.59-20.27), with adjusted HR 0.60 (95%CI: 0.45-0.79, p&amp;lt;0.001). Unlike HR+HER2- disease, cumulative hazard of relapse or death in yr 5-10 in HR+HER2+ pts treated with TH was lower at 8.6% (95%CI 6.16-10.97). This is slightly higher than pts with HR-HER2+ at 5.75% (95%CI 3.18-8.24, adjusted HR=1.7, 95%CI 1.01-2.85, p=0.046). Compared to 22% and 31% risk of relapse after 5 yr in N0 and N1 HR+HER2- in recent publication, 3.23% (95%CI 0-9.25) of pts with HR+HER2+ with N0 developed recurrence in yr 5-10 and 6.39% (95%CI 3.09-9.59) in those with N1 disease. Albeit rare, pts with Luminal A tumors had 89.8% relapse-free survival at 10 yr compared to Luminal B 82.3%, HER2 enriched 76.8%, and Basal 74.19%. Conclusions: Even without extended adjuvant endocrine therapy in both trials, risk of late relapse in HR+ HER2+ appeared to be low, particularly in pts with N0 or N1 disease. The benefit of extended adjuvant endocrine therapy in this group of TH pts, particularly those with no lymph node involvement, may not outweigh side effects of continuing endocrine therapy beyond 5 yr. Support: U10CA180821, U10CA18082, U24CA196171; U10CA180868, UG1CA189867, U10CA180822, U24CA196067; ClinicalTrials.gov Id: NCT00005970, NCT00004067 Cumulative probability (%) of relapse or death.Nodal statusYear 0-5Year 5-10* HR+HR-HR+HR- Cumulative hazard (95%CI)Cumulative hazard (95%CI)Cumulative hazard (95%CI)Cumulative hazard (95%CI)07.71 ( 0.00, 15.74)16.05 ( 7.98, 23.42)3.23 ( 0.00, 9.25)8.86 ( 1.05, 16.04)16.70 ( 3.66, 9.64)10.63 ( 5.99, 15.04)6.39 ( 3.09, 9.59)5.57 ( 1.46, 9.51)213.68 ( 7.42, 19.52)20.99 (12.98, 28.26)5.10 ( 0.64, 9.35)1.22 ( 0.00, 3.57)310.25 ( 2.76, 17.17)39.76 (24.11, 52.18)13.19 ( 3.54, 21.88)8.52 ( 0.00, 19.16)*Based on pts who survived 5 yr without relapse Citation Format: Chumsri S, Li Z, Serie DJ, Mashadi-Hossein A, Colon-Otero G, Song N, Pogue-Geile K, Gavin P, Paik S, Moreno-Aspitia A, Perez EA, Thompson EA. Incidence of late relapse in HER2-positive (HER2+) breast cancer patients receiving adjuvant trastuzumab: Combined analysis of NCCTG (Alliance) N9831 and NSABP (NRG) B31 [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 PD3-02.

Abstract OT1-05-02: 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 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) &amp;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&amp;lt;30mm58 &amp;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.

Comprehensive clinical and molecular analyses of neuroendocrine carcinomas of the breast

Neuroendocrine breast carcinomas represent a rare subtype of breast cancer. Their definition, prevalence, and prognosis remain controversial in the literature. The 2012 WHO classification of breast cancer categorizes neuroendocrine carcinomas into three morphologically distinct subtypes: well-differentiated neuroendocrine tumors, poorly differentiated neuroendocrine carcinomas, and invasive breast carcinomas with neuroendocrine differentiation. We aimed to gain insight into the clinical, morphologic, phenotypic, and molecular features of 47 neuroendocrine breast carcinomas. Targeted next-generation sequencing by an AmpliSeq 22 cancer gene hotspot panel and the Prosigna assay were performed on 42/47 and 35/47 cases, respectively. Average age at diagnosis was 69 years. All tumors were estrogen receptor-positive and the large majority expressed progesterone receptor (89%), GATA3 (98%), FOXA1 (96%), and CK8/18 (98%). There was an almost equal distribution of luminal A (52%) and B (48%) carcinomas. Almost half of the cohort (49%) displayed a high risk of recurrence score with the Prosigna test. Patients with a neuroendocrine carcinoma had a shorter disease-free survival compared with those affected by carcinomas of no special type matched for age, size, grade, and estrogen receptor status. No significant differences were observed in terms of overall survival. Stratification of neuroendocrine carcinomas using the 2012 WHO criteria did not reveal statistically significant differences among the distinct categories (well-differentiated neuroendocrine tumors, poorly differentiated neuroendocrine carcinomas, and invasive breast carcinomas with neuroendocrine differentiation), in terms of either progression-free or overall survival. Our targeted sequencing analysis found three cases (7%) harboring a PIK3CA mutation, and in three other cases (7%) TP53 mutations were detected. This study showed that neuroendocrine breast carcinoma is a distinct subtype of luminal carcinoma with a low rate of PIK3CA mutations and with an aggressive clinical behavior. An accurate identification of neuroendocrine differentiation may be useful to better tailor patient adjuvant therapy within luminal carcinomas.

Prospective, multicenter French study evaluating the clinical impact of the Breast Cancer Intrinsic Subtype-Prosigna® Test in the management of early-stage breast cancers.

PurposeThe Prosigna® breast cancer prognostic gene signature assay identifies a gene-expression profile that permits the classification of tumors into subtypes and gives a score for the risk of recurrence (ROR) at 10 years. The primary objective of this multicenter study was to evaluate the impact of Prosigna’s assay information on physicians’ adjuvant treatment decisions in patients with early-stage breast cancer. Secondary objectives were to assess confidence of practitioners in their therapeutic recommendations before and after the added information provided by the Prosigna assay; and to evaluate the emotional state of patients before and after the Prosigna test results.MethodsConsecutive patients with invasive early-stage breast cancer were enrolled in a prospective, observational, multicenter study carried out in 8 hospitals in France. The Prosigna test was carried out on surgical specimens using the nCounter® Analysis System located at the Institut Curie. Both before and after receiving the Prosigna test results, physicians completed treatment confidence questionnaires and patients completed questionnaires concerning their state of anxiety, the difficulties felt in face of the therapy and quality of life. Information was also collected at 6 months regarding the physicians’ opinion on the test results and the patients’ degree of anxiety, difficulties with therapy and quality of life.ResultsBetween March 2015 and January 2016, 8 study centers in France consecutively enrolled 210 postmenopausal women with estrogen receptor (ER) positive, human epidermal growth hormone-2 (HER-2) negative, and node negative tumors, either stage 1 or stage 2. Intrinsic tumor subtypes as assessed by the Prosigna test were 114 (58.2%) Luminal A, 79 (40.3%) Luminal B, 1 (0.5%) HER-2 enriched (HER-2E), and 2 (1.0%) basal-like. Before receiving the Prosigna test results, physicians categorized tumor subtypes based on immunohistochemistry (IHC) as Luminal A in 126 (64%) patients and Luminal B in 70 (36%) patients, an overall discordance rate of 25%. The availability of Prosigna assay results was significantly associated with the likelihood of change in treatment recommendations, with 34 patients (18%) having their treatment plan changed from Adjuvant Chemotherapy to No Adjuvant Chemotherapy or vice versa (p<0.001, Fisher’s exact test). Prosigna test results also decreased patients’ anxiety about the chosen adjuvant therapy, and improved emotional well-being and measures of personal perceptions of uncertainty.ConclusionsThe results of this prospective decision impact study are consistent with 2 previous, identically designed studies carried out in Spain and Germany. The availability of Prosigna test results increased the confidence of treating physicians in their adjuvant treatment decisions, and led to an 18% change in chemotherapy treatment plan (from Adjuvant Chemotherapy to No Adjuvant Chemotherapy or vice versa). Prosigna testing decreased anxiety and improved measures of health-related quality of life in patients facing adjuvant therapy. The 25% discordance between Prosigna test and IHC subtyping underlines the importance of molecular testing for optimal systemic therapy indications in early breast cancer.

Impact of the Prosigna (PAM50) assay on adjuvant clinical decision making in patients with early stage breast cancer: Results of a prospective multicenter public program.

e12062 Background: Prosigna is a standardized test based on the PAM50 gene signature which provides information on intrinsic subtype and risk of recurrence (ROR) score predicting 10y recurrence probability (NanoString Technologies, Inc., Seattle, WA). We evaluated Prosigna’s impact on adjuvant treatment decision beyond immunohistochemistry (IHC) testing. Methods: 125 pre- and postmenopausal EBC patients with ER+, HER2-, pT1-T2 pN0/pN1mi were enrolled (12 centers; 8/2015-11/2016). FFPE specimens were centrally analyzed using Prosigna to classify patients according to the intrinsic tumor subtype and ROR score. The primary endpoint was the impact of the Prosigna test on adjuvant treatment decision. Results: 64% of tumors were classified by PAM50 as Lum A, 35% as Lum B, 0% Basal, 1% HER2-E. The intrinsic subtype concordance between immunohistochemistry (IHC) and Prosigna (n = 119) was 61.3%. In Lum B tumors by IHC the discordance rate was the highest (48% were reclassified as Lum A by Prosigna) as shown in the Table. ROR risk groups were as follows: ROR low (54; 43%), ROR interm. (38; 30%), and ROR high (33; 27%). Prosigna results led to a treatment recommendation change in 49 (39%) patients. In the 76 (61%) cases with the initial recommendation of Hormonal Therapy (HT) alone the final decision changed to chemotherapy (CT)+HT (CHT) in 24 (32%) patients. In the 49 (39%) cases in which the initial recommendation was CHT the final decision changed to HT in 25 (51%) patients. Among the 38 patients with Prosigna intermediate risk, 21 (55%) were allocated to HT. Conclusions: In this prospective decision impact study, Prosigna results led to a 39% change in adjuvant therapy indication. Patients with initial indication of CHT were changed to HT alone in &gt; 50% of cases. Thus, Prosigna results influenced the treatment decisions and reinforced its clinical utility in real-world settings. The intrinsic subtype classification based on IHC didn’t show to be an adequate surrogate for the genomic subtypes as determined by Prosigna. [Table: see text]

The West German Study Group Breast Cancer Intrinsic Subtype study: a prospective multicenter decision impact study utilizing the Prosigna assay for adjuvant treatment decision-making in estrogen-receptor-positive, HER2-negative early-stage breast cancer

Purpose: The West German Study Group (WSG) Breast Cancer Intrinsic Subtype (BCIST) study was designed to assess the influence of Prosigna gene signature assay results on physicians’ adjuvant treatment recommendations by determining the extent of change in pre-test treatment recommendations following assay results. Secondary objectives were to assess the influence of Prosigna results on physicians’ confidence in their therapeutic recommendations and on patients’ decisional conflict status, anxiety levels, and functional status. Methods: This prospective, observational, decision impact study enrolled consecutive postmenopausal patients with estrogen-receptor (ER)-positive, HER2-negative, lymph-node-negative early-stage breast cancer in 11 centers in Germany. Physicians based their pre-test adjuvant treatment recommendations on standard clinico-pathological parameters. Tumor specimens were assayed using the Prosigna test in a WSG central pathology laboratory following manufacturer’s guidelines. An independent pathology laboratory performed subsequent Prosigna assays on tumor sections to assess assay result concordance with the central laboratory. Physicians completed treatment confidence questionnaires prior to and after receiving Prosigna test results. Patients completed standardized questionnaires on decisional conflict, anxiety, and health status both before and after Prosigna testing. Results: The present study population consisted predominantly of low-to-intermediate risk patients (N = 198). Prosigna had 29.3% discordance in intrinsic subtyping with local immunohistochemistry test results. After Prosigna test results, a change in the adjuvant therapy recommendation occurred in 36 (18.2%) patients; 22 (11.1%) patients switched from no chemotherapy to chemotherapy. After Prosigna test results, physicians expressed increased confidence in their prognostic assessment in 87.9% of patients, and increased confidence in their treatment recommendation in 89.4%. Patients reported improved anxiety and emotional/functional well-being after receiving Prosigna test results. Conclusions: Use of the Prosigna assay led to a change in 18.2% of adjuvant treatment decisions. Prosigna testing was associated with increased patient and physician confidence in treatment decisions, and with decreased patient anxiety and improved well-being. Any comparison of the therapeutic decision-making impacts of different genomic assays must account for potential confounding factors.

Development and verification of the PAM50-based Prosigna breast cancer gene signature assay.

BackgroundThe four intrinsic subtypes of breast cancer, defined by differential expression of 50 genes (PAM50), have been shown to be predictive of risk of recurrence and benefit of hormonal therapy and chemotherapy. Here we describe the development of Prosigna™, a PAM50-based subtype classifier and risk model on the NanoString nCounter Dx Analysis System intended for decentralized testing in clinical laboratories.Methods514 formalin-fixed, paraffin-embedded (FFPE) breast cancer patient samples were used to train prototypical centroids for each of the intrinsic subtypes of breast cancer on the NanoString platform. Hierarchical cluster analysis of gene expression data was used to identify the prototypical centroids defined in previous PAM50 algorithm training exercises. 304 FFPE patient samples from a well annotated clinical cohort in the absence of adjuvant systemic therapy were then used to train a subtype-based risk model (i.e. Prosigna ROR score). 232 samples from a tamoxifen-treated patient cohort were used to verify the prognostic accuracy of the algorithm prior to initiating clinical validation studies.ResultsThe gene expression profiles of each of the four Prosigna subtype centroids were consistent with those previously published using the PCR-based PAM50 method. Similar to previously published classifiers, tumor samples classified as Luminal A by Prosigna had the best prognosis compared to samples classified as one of the three higher-risk tumor subtypes. The Prosigna Risk of Recurrence (ROR) score model was verified to be significantly associated with prognosis as a continuous variable and to add significant information over both commonly available IHC markers and Adjuvant! Online.ConclusionsThe results from the training and verification data sets show that the FDA-cleared and CE marked Prosigna test provides an accurate estimate of the risk of distant recurrence in hormone receptor positive breast cancer and is also capable of identifying a tumor's intrinsic subtype that is consistent with the previously published PCR-based PAM50 assay. Subsequent analytical and clinical validation studies confirm the clinical accuracy and technical precision of the Prosigna PAM50 assay in a decentralized setting.Electronic supplementary materialThe online version of this article (doi:10.1186/s12920-015-0129-6) contains supplementary material, which is available to authorized users.