Association of Genomic Prostate Score at positive margin with recurrence after radical prostatectomy.

OBJECTIVES: To evaluate the utility of the 17-gene Genomic Prostate Score® (GPS; MDxHealth, Irvine, CA, USA) performed on prostate cancer at the positive margin of the radical prostatectomy (RP) for its association with risk of subsequent biochemical recurrence (BCR). PATIENTS AND METHODS: We designed a case-cohort for the outcome of BCR, selecting 223 from a cohort of 813 RP patients treated at Johns Hopkins from 2008 to 2017 with positive margins and available clinical data; of these, 213 had available tissue and clinical data. RNA was isolated from formalin-fixed paraffin-embedded tumour tissue adjacent to the positive surgical margin and the GPS was evaluable in 203 of these patients with a score ranging from 0 to 100, with higher scores indicating higher risk. All patients underwent RP with or without adjuvant radiation therapy (ART). The statistical analysis employed Cox proportional hazards regression models for outcome of BCR weighted for case-cohort design. RESULTS: In univariable analysis, every 20-unit increase in the GPS was associated with a nearly threefold increase in risk of BCR (hazard ratio [HR] per 20 units 2.82, P < 0.001). In a multivariable Cox model adjusted for age, race, Cancer of the Prostate Risk Assessment Postsurgical score, Grade Group at the positive margin, and ART, the GPS was significantly associated with BCR (HR 1.56 per 20 units; 95% confidence interval 1.11-2.19; P = 0.011). The study is limited by its retrospective and single institution design. CONCLUSIONS: The GPS at the positive surgical margin could help stratify prognosis and inform clinical decision-making regarding adjuvant therapy after RP.

Clinical and Genomic Risk for Late Breast Cancer Recurrence and Survival.

BACKGROUND: The 21-gene recurrence score (RS) assay (Oncotype DX) is used to guide adjuvant chemotherapy use for patients with hormone receptor-positive, HER2 (human epidermal growth factor receptor 2)-negative, axillary node-negative breast cancer. Its role, however, in providing prognostic information for late distant recurrence when added to clinicopathologic prognostic factors is unknown. METHODS: A patient-specific meta-analysis including 10,004 women enrolled in three trials was updated using extended follow-up data from TAILORx, integrating the RS with histologic grade, tumor size, and age at surgery for the RSClin tool. Cox models integrating clinicopathologic factors and the RS were compared by using likelihood ratio (LR) tests. External validation of prognosis for distant recurrence in years 0 to 10 and 5 to 10 was performed in an independent cohort of 1098 women in a real-world registry. RESULTS: RSClin provided significantly more prognostic information than either the clinicopathologic factors (ΔLR chi-square, 86.2; P<0.001) or RS alone (ΔLR chi-square, 131.0; P<0.001). The model was prognostic in an independent cohort for distant recurrence by 10 years after diagnosis (standardized hazard ratio, 1.56; 95% confidence interval, 1.25 to 1.94), was associated with late distant recurrence risk between 5 and 10 years after diagnosis (standardized hazard ratio, 1.78; 95% confidence interval, 1.25 to 2.55), and approximated the observed 10-year distant recurrence risk (Lin concordance, 0.87) and 5- to 10-year distant recurrence risk (Lin concordance, 0.92). CONCLUSIONS: The 21-gene RS is prognostic for distant recurrence and overall survival in early breast cancer. A model integrating the 21-gene RS and clinicopathologic factors improved estimates of distant recurrence risk compared with either used individually and stratified late distant recurrence risk. (Funded by the National Cancer Institute, National Institutes of Health [U10CA180820, U10CA180794, UG1CA189859, U10CA180868, and U10CA180822] and others.).

Advancing Evidence Generation for Circulating Tumor DNA: Lessons Learned from A Multi-Assay Study of Baseline Circulating Tumor DNA Levels across Cancer Types and Stages.

Circulating tumor DNA (ctDNA) holds promise as a biomarker for predicting clinical responses to therapy in solid tumors, and multiple ctDNA assays are in development. However, the heterogeneity in ctDNA levels prior to treatment (baseline) across different cancer types and stages and across ctDNA assays has not been widely studied. Friends of Cancer Research formed a collaboration across multiple commercial ctDNA assay developers to assess baseline ctDNA levels across five cancer types in early- and late-stage disease. This retrospective study included eight commercial ctDNA assay developers providing summary-level de-identified data for patients with non-small cell lung cancer (NSCLC), bladder, breast, prostate, and head and neck squamous cell carcinoma following a common analysis protocol. Baseline ctDNA levels across late-stage cancer types were similarly detected, highlighting the potential use of ctDNA as a biomarker in these cancer types. Variability was observed in ctDNA levels across assays in early-stage NSCLC, indicative of the contribution of assay analytical performance and methodology on variability. We identified key data elements, including assay characteristics and clinicopathological metadata, that need to be standardized for future meta-analyses across multiple assays. This work facilitates evidence generation opportunities to support the use of ctDNA as a biomarker for clinical response.

BRCA1/2 mutation carriers vs the general breast cancer population (N = 799,986): 21-gene assay-based molecular characterization.

PURPOSE: We compared 21-gene recurrence score (RS) distribution and expression of the single-gene/gene groups within this assay between BC patients with pathogenic variants (PV) in BRCA1/2 vs the general 21-gene-tested BC population. METHODS: This retrospective study included consecutive 21-gene-tested female ER + HER2-negative BC patients with germline PVs in BRCA1/2. RS/gene expression data were compared to a previously described commercial use database (CDB, N = 799,986). Chi-square and 1-sample t test were used to compare RS distribution and single-gene/gene group scores between the study group and the CDB. RESULTS: Study group patients (N = 81) were younger and their RS results were higher compared to the CDB (age: median [IQR], 56 [47-61.5] vs 60 [51-67] years; p < 0.001; proportion of patients with RS ≥ 26: 49.4% vs 16.4%, p < 0.001). Expression of 12/16 cancer genes in the assay and the ER, proliferation, and invasion gene group scores differed significantly between the study group and the CDB, all in a direction contributing to higher RS. The differences between the study group and the CDB were mostly retained, upon stratifying the patients by menopausal status. CONCLUSION: BC patients with PVs in BRCA1/2 have higher RS results that stem from distinct gene expression profiles in the majority of genes in the 21-gene assay.

Gene Expression and Benefit of Chemotherapy in Women With Node-Negative, Estrogen Receptor-Positive Breast Cancer.

PURPOSE: The 21-gene recurrence score (RS) assay quantifies the likelihood of distant recurrence in women with estrogen receptor-positive, lymph node-negative breast cancer treated with adjuvant tamoxifen. The relationship between the RS and chemotherapy benefit is not known. METHODS: The RS was measured in tumors from the tamoxifen-treated and tamoxifen plus chemotherapy-treated patients in the National Surgical Adjuvant Breast and Bowel Project (NSABP) B20 trial. Cox proportional hazards models were utilized to test for interaction between chemotherapy treatment and the RS. RESULTS: A total of 651 patients were assessable (227 randomly assigned to tamoxifen and 424 randomly assigned to tamoxifen plus chemotherapy). The test for interaction between chemotherapy treatment and RS was statistically significant (P = .038). Patients with high-RS (≥ 31) tumors (ie, high risk of recurrence) had a large benefit from chemotherapy (relative risk, 0.26; 95% CI, 0.13 to 0.53; absolute decrease in 10-year distant recurrence rate: mean, 27.6%; SE, 8.0%). Patients with low-RS (< 18) tumors derived minimal, if any, benefit from chemotherapy treatment (relative risk, 1.31; 95% CI, 0.46 to 3.78; absolute decrease in distant recurrence rate at 10 years: mean, -1.1%; SE, 2.2%). Patients with intermediate-RS tumors did not appear to have a large benefit, but the uncertainty in the estimate can not exclude a clinically important benefit. CONCLUSION: The RS assay not only quantifies the likelihood of breast cancer recurrence in women with node-negative, estrogen receptor-positive breast cancer, but also predicts the magnitude of chemotherapy benefit.

Ultrasensitive Circulating Tumor DNA Pilot Study Distinguishes Complete Response and Partial Response With Immunotherapy in Patients With Metastatic Renal Cell Carcinoma.

PURPOSE: Circulating tumor DNA (ctDNA) has been validated across multiple indications in the adjuvant and surveillance settings. We evaluated whether targeted digital sequencing (TARDIS) may distinguish a partial response (PR) from a complete response (CR) among patients with metastatic renal cell carcinoma (mRCC) receiving immune checkpoint inhibitor (ICI) therapy. MATERIALS AND METHODS: Eligible patients had mRCC that yielded a PR or CR to ICI therapy. Peripheral blood was obtained at a single time point for ctDNA analysis. TARDIS was used for quantification of average variant allele fractions (VAFs). Our primary objective was to determine the association between VAFs and depth of response (PR v CR). A secondary objective was to determine whether VAFs were associated with disease progression. RESULTS: Twelve patients were analyzed, nine of whom achieved a PR (75%). Patients received either nivolumab monotherapy (50%) or nivolumab plus ipilimumab (50%). ctDNA analysis incorporated an average of 30 patient-specific mutations (range, 19-35); average coverage depth was 103,342 reads per target. TARDIS quantified a significant difference in VAFs between PR and CR (median, 0.181% [IQR, 0.077%-0.420%] v 0.007% [IQR, 0.0%-0.028%], respectively [P = .014]). Of the 12 patients in the series, six patients demonstrated radiographic progression subsequent to ctDNA assessment. Patients who progressed on subsequent scans had significantly higher ctDNA than those who maintained their response (median, 0.362% [IQR, 0.181%-2.71%] v 0.033% [IQR, 0.007%-0.077%], respectively [P = .026]). CONCLUSION: In this pilot study, TARDIS accurately differentiated PR from CR among patients with mRCC receiving immunotherapy, and also prospectively identified patients at risk for subsequent progression. Given these findings, we envision subsequent studies that validate these results and investigate the utility of this assay to discern appropriate candidates for discontinuation of immunotherapy.

Population-based estimate for the correlation of the Oncotype Dx Breast Recurrence Score® result and Ki-67 IHC MIB-1 pharmDx in HR+, HER2-, node-positive early breast cancer.

BACKGROUND: The United States Food and Drug Administration recently approved a Ki-67 immunohistochemistry (IHC) assay to identify patients with early breast cancer at high disease recurrence risk. The Oncotype Dx Breast Recurrence Score® assay has been validated in hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) invasive breast cancer (IBC) to predict chemotherapy benefit and distant recurrence risk, regardless of nodal status. This study assessed the correlation between Recurrence Score® (RS) results and the Ki-67 IHC MIB-1 pharmDx assay. METHODS: HR+, HER2-, N1 IBC samples with RS results were examined by Ki-67 IHC; 311 specimens were collected, including 275 without regard to RS ("unselected RS") and 36 more with RS 26-100; 12 were lymph node negative upon pathology report review, and one had no Ki-67 score, leaving 262 unselected RS and 298 total samples. Spearman rank correlation was calculated using the unselected samples and a weighted rank correlation using all samples. A receiver operating characteristic (ROC) curve for predicting high RS (26-100) from Ki-67 was constructed. RESULTS: The Spearman rank correlation between Ki-67 and RS results was moderately positive (unselected RS samples: 0.396; 95% confidence interval [CI] 0.288-0.493; all samples: 0.394; 95% CI 0.294-0.486). While 71% of samples with RS 26-100 had Ki-67 ≥ 20%, 75% with RS 0-25 had Ki-67 < 20%. ROC area under the curve was 0.792 (95% CI 0.725-0.859). CONCLUSIONS: The moderately positive correlation is consistent with previous analyses suggesting the Oncotype Dx® assay and Ki-67 IHC MIB-1 assay should not be used interchangeably in clinical practice.

Patient-specific meta-analysis of 12-gene colon cancer recurrence score validation studies for recurrence risk assessment after surgery with or without 5FU and oxaliplatin.

Background: Individualized estimates of the risk of recurrence in colon cancer patients are needed that reflect current medical practice and available treatment options. Methods: Three validation studies of the 12-gene colon recurrence score assay were used with pre-specified patient-specific meta-analysis (PSMA) methods to integrate the 12-gene Oncotype DX Colon Recurrence Score result (RS) with the clinical and pathology risk factors stage, T-stage, mis-match repair (MMR) status, and number of nodes examined to calculate individualized recurrence risk estimates. Baseline risk estimation used the most recent studies, so the risk estimates reflect current medical practice. The effect of fluorouracil (5FU) was estimated with a meta-analysis of two studies. The effect of oxaliplatin was estimated using one of the RS assay validation studies, in which patients were randomized to 5FU with or without oxaliplatin. Results: The RS result and each of the clinical-pathologic factors provided independent prognostic information for recurrence. Among stage II, T3, MMR-proficient patients with ≥12 nodes examined (the most common scenario), patients with RS ≤30 (approximately 48%) have estimated 5-year recurrence risk ≤10% with surgery alone. Among stage IIIA/B, T3, MMR-deficient patients with ≥12 nodes examined, patients with RS ≤19 (approximately 14%) have an estimated 5-year recurrence risk ≤10% with surgery alone. Among stage IIIA/B, T3, MMR-proficient patients with ≥12 nodes examined, those with RS ≤14 (approximately 6%) have estimated 5-year recurrence risk ≤10% with 5FU alone. Discussion: The PSMA integrates the 12-gene colon RS result with clinical and pathology factors to provide individualized recurrence risk estimates that reflect current medical practice. The risk estimates are in a range that may help inform treatment decisions for a substantial number of stage II and stage III patients.

Validating the association of adverse pathology with distant metastasis and prostate cancer mortality 20-years after radical prostatectomy.

PURPOSE: To assess the association of adverse pathology (AP), defined as high-grade (≥ Gleason Grade Group 3) and/or non-organ confined disease, with long-term oncologic outcomes after radical prostatectomy (RP). MATERIALS AND METHODS: Using a stratified cohort sampling design, we evaluated the association of AP with the risk of distant metastasis (DM) and prostate cancer-specific mortality (PCSM) up to 20 years after RP in 428 patients treated between 1987 to 2004. Cox regression of cause-specific hazards was used to estimate the absolute risk of both endpoints, with death from other causes treated as a competing risk. Additionally, subgroup analysis in patients with low and/or intermediate-risk disease, who are potentially eligible for active surveillance (AS), was performed. RESULTS: Within the cohort sample, 53% of men exhibited AP at time of RP, with median follow up of 15.5 years (IQR 14.6-16.6 years) thereafter. Adverse pathology was highly associated with DM and PCSM in the overall cohort (HR 12.30, 95% confidence interval [CI] 5.30-28.55, and HR 10.03, 95% CI 3.42-29.47, respectively, both P < 0.001). Adverse pathology was also highly associated with DM and PCSM in the low/intermediate-risk subgroup (HR 10.48, 95% CI 4.18-26.28, and 8.60, 95% CI 2.40-30.48, respectively, both P < 0.001). CONCLUSIONS: Adverse pathology at the time of RP is highly associated with future development of DM and PCSM. Accurate prediction of AP may thus be useful for individualizing risk-based surveillance and treatment strategies.

A Comparison of Predicted Ipsilateral Tumor Recurrence Risks in Patients With Ductal Carcinoma in Situ of the Breast After Breast-Conserving Surgery by Breast Radiation Oncologists, the Van Nuys Prognostic Index, the Memorial Sloan Kettering Cancer Center DCIS Nomogram, and the 12-Gene DCIS Score Assay.

PURPOSE: To compare ipsilateral breast event (IBE) risks in patients with ductal carcinoma in situ of the breast (DCIS) post-lumpectomy, as estimated by breast radiation oncologists, the Van Nuys Prognostic Index, the Memorial Sloan Kettering Cancer Center (MSKCC) DCIS nomogram, and the 12-gene Oncotype DX DCIS score assay. METHODS AND MATERIALS: Consecutive DCIS cases treated with lumpectomy from November 2011 to August 2014 with available DCIS score results were identified. Three radiation oncologists independently estimated the 10-year IBE risk. The Van Nuys Prognostic Index and MSKCC nomogram 10-year IBE risk estimates were generated. Differences and correlations between the IBE estimates and clinicopathologic factors were evaluated. RESULTS: Ninety-one patients were identified for inclusion. Forty-eight percent would have been ineligible for the E5194 study. The mean risk of IBE from the DCIS score assay was 12.4%, compared with a range of 18.9% to 26.8% from other sources. The mean IBE risk from the DCIS score assay was lower regardless of E5194 eligibility. The MSKCC nomogram and DCIS score assay risk estimates were weakly correlated with each other (P = .23) and were each moderately correlated with the other risk estimates (P = .41-.56). When applying the radiation oncologists' treatment recommendations based on their proposed risk cutoffs, evaluating risk according to the DCIS score assay led to the highest proportion of patients recommended excision alone. CONCLUSIONS: IBE risk estimates for this general community cohort of DCIS cases vary significantly among commonly available clinical predictive tools and individual radiation oncologist estimates. Surgical margins and tumor size continue to factor prominently in radiation oncologist decision algorithms. The differences found between the IBE risk estimate methods suggests that they are not interchangeable and the methods that rely on clinicopathologic features may tend to overestimate risk.

Molecular Drivers of Oncotype DX, Prosigna, EndoPredict, and the Breast Cancer Index: A TransATAC Study.

PURPOSE: The Oncotype DX Recurrence Score (RS), Prosigna Prediction Analysis of Microarray 50 (PAM50) Risk of Recurrence (ROR), EndoPredict (EP), and Breast Cancer Index (BCI) are used clinically for estimating risk of distant recurrence for patients receiving endocrine therapy. Discordances in estimates occur between them. We aimed to identify the molecular features that drive the tests and lead to these differences. PATIENTS AND METHODS: Analyses for RS, ROR, EP, and BCI were conducted by the manufacturers in the TransATAC sample collection that consisted of the tamoxifen or anastrozole arms of the ATAC trial. Estrogen receptor-positive/human epidermal growth factor receptor 2 (HER2)-negative cases without chemotherapy treatment were included in which all four tests were available (n = 785). Clinicopathologic features included in some tests were excluded from the comparisons. Estrogen, proliferation, invasion, and HER2 module scores from RS were used to characterize the respective molecular features. Spearman correlation and analysis of variance tests were applied. RESULTS: There were moderate to strong correlations among the four molecular scores (ρ = 0.63-0.74) except for RS versus ROR (ρ = 0.32) and RS versus BCI (ρ = 0.35). RS had strong negative correlation with its estrogen module (ρ = -0.79) and moderate positive correlation with its proliferation module (ρ = 0.36). RS's proliferation module explained 72.5% of ROR's variance, while the estrogen module explained only 0.6%. Most of EP's and BCI's variation was accounted for by the proliferation module (50.0% and 54.3%, respectively) and much less by the estrogen module (20.2% and 2.7%, respectively). CONCLUSION: In contrast to common understanding, RSs are determined more strongly by estrogen-related features and only weakly by proliferation markers. However, the EP, BCI, and particularly ROR scores are determined largely by proliferative features. These relationships help to explain the differences in the prognostic performance of the tests.

The run-in phase of the prospective WSG-ADAPT HR+/HER2- trial demonstrates the feasibility of a study design combining static and dynamic biomarker assessments for individualized therapy in early breast cancer.

BACKGROUND: Endocrine sensitivity, as determined by response of the proliferation marker Ki-67 to short-term preoperative endocrine therapy (ET), is currently not included in adjuvant treatment decisions in hormone receptor (HR)+/human epidermal growth factor receptor 2 (HER2)- breast cancer (BC). METHODS: The prospective WSG-ADAPT HR+/HER2- trial included patients with N0/N1 early BC who were candidates for adjuvant chemotherapy based on clinical-pathological criteria alone. The trial utilized a genomic assessment [the Recurrence Score (RS)] plus endocrine sensitivity testing to guide treatment. All patients received 3 (±1) weeks of preoperative induction ET. According to protocol, patients with RS 0-11 or RS 12-25 plus endocrine proliferation response (EPR, post-induction Ki-67 ⩽ 10%) were to be spared adjuvant chemotherapy. RESULTS: The ADAPT HR+/HER2- trial run-in phase included 407 patients with baseline RS, of whom 386 (median age: 54 years) had complete data for Ki-67 at both baseline and post-induction. RS distribution: 23.1% RS 0-11, 58.3% RS 12-25, and 18.7% RS 26-100. EPR occurred in 84.3%, 76.0%, and 36.1% of these RS groups, respectively. Differences in EPR proportions (RS 26-100 versus others, RS 0-11 versus others) were significant (both p < 0.001); Ki-67 quotients were higher for RS 26-100 (p = 0.02, Mann-Whitney). In premenopausal women (n = 146, mostly tamoxifen-treated), median quotient of Ki-67 level (post/pre) was significantly higher than in postmenopausal women (n = 222, mostly aromatase-inhibitor treated; 0.67 versus 0.25, p < 0.001). EPR was significantly associated with baseline estrogen-receptor status as determined by immunohistochemistry (p = 0.002) or real-time polymerase chain reaction (p < 0.001). Also, a strong correlation was observed between RS measured pre- and post-ET (RS = 0.7, n = 181). CONCLUSIONS: This phase of the WSG-ADAPT HR+/HER2- trial confirms trial design estimates of RS and EPR. It indicates that the ADAPT concept of combining static and dynamic biomarker assessment for individualized therapy decisions in early BC is feasible using the EPR criterion post-induction Ki-67 ⩽ 10%. CLINICALTRIALSGOV IDENTIFIER: NCT01779206.

Molecular characterization of breast cancer needle core biopsy specimens by the 21-gene Breast Recurrence Score test.

BACKGROUND AND OBJECTIVE: Recent COVID-19 pandemic guidelines recommend genomic assessment of core biopsies to help guide treatment decisions in estrogen receptor (ER)-positive early-stage breast cancer. Herein we characterize biopsy and excisional breast cancer specimens submitted for 21-gene testing. METHODS: US samples submitted to Genomic Health for 21-gene testing (01/2004-04/2020) were assessed by pathologists and analyzed by a standardized quantitative reverse transcription-polymerase chain reaction. Predefined cutoffs were: ESR1 (positive ≥6.5), PGR (positive ≥5.5), and ERBB2 (negative <10.7). ER status by immunohistochemistry (IHC) and lymph node status were determined locally. Median and interquartile range were reported for continuous variables, and total and percent for categorical variables. Distributions were assessed overall, by age, and by nodal involvement. RESULTS: Of 919 701 samples analyzed, 13% were biopsies and 87% were excisions. Initial assay success rates were 94.5% (biopsies) and 97.3% (excisions). ER IHC concordance with central ESR1 was 96.8% (biopsies) and 97.6% (excisions). Biopsy and excisional medians were: Recurrence Score results 16 (each); ESR1 10.2 (each); PGR 7.7 and 7.6; ERBB2 9.4 and 9.2, respectively. CONCLUSIONS: Biopsy submissions for 21-gene testing are common and consistently generate results that are very similar to the experience with excisions. The 21-gene test can be performed reliably on core biopsies.

Genomic comparison of paired primary breast carcinomas and lymph node macrometastases using the Oncotype DX Breast Recurrence Score® test.

PURPOSE: Adjuvant therapy decisions may in part be based on results of Oncotype DX Breast Recurrence Score® (RS) testing of primary tumors. When necessary, lymph node metastases may be considered as a surrogate. Here we evaluate the concordance in gene expression between primary breast cancers and synchronous lymph node metastases, based on results from quantitative RT-PCR-based RS testing between matched primary tumors and synchronous nodal metastases. METHODS: This retrospective, exploratory study included patients (≥ 18 years old) treated at our center (2005-2009) who had ER+ , HER2-negative invasive breast cancer and synchronous nodal metastases with available tumor blocks from both sites. Paired tissue blocks underwent RS testing, and RS and single-gene results for ER, PR, and HER2 were explored between paired samples. RESULTS: A wide distribution of RS results in tumors and in synchronous nodal metastases were modestly correlated between 84 paired samples analyzed (Pearson correlation 0.69 [95% CI 0.55-0.78]). Overall concordance in RS group classification between samples was 63%. ER, PR, and HER2 by RT-PCR between the primary tumor and lymph node were also modestly correlated (Pearson correlation [95% CI] 0.64 [0.50-0.75], 0.64 [0.49-0.75], and 0.51 [0.33-0.65], respectively). Categorical concordance (positive or negative) was 100% for ER, 77% for PR, and 100% for HER2. CONCLUSIONS: There is modest correlation in continuous gene expression, as measured by the RS and single-gene results for ER, PR, and HER2 between paired primary tumors and synchronous nodal metastases. RS testing for ER+ breast cancer should continue to be based on analysis of primary tumors.

Accurate Estrogen Receptor Quantification in Patients with Negative and Low-Positive Estrogen-Receptor-Expressing Breast Tumors: Sub-Analyses of Data from Two Clinical Studies.

INTRODUCTION: Accurate assessment of estrogen receptor (ER) expression is crucial to ensure that patients with early breast cancer are accurately identified for appropriate treatment with endocrine therapy. Reverse transcriptase polymerase chain reaction (RT-PCR), compared with immunohistochemistry (IHC), may provide a more precise indication of ER status. Data were pooled and analyzed from two independent, but similarly designed, studies that examined ER status by IHC and the 21-gene Recurrence Score that employs RT-PCR-based methodology. METHODS: Tumor tissue from patients with early stage breast cancer where ER status could be determined by both IHC and RT-PCR was included. ER status by IHC staining was defined as ER-negative (< 1%), ER-low+ (1-10%), or ER+ (> 10%). ER status by RT-PCR was defined as ER-negative (≤ 6.5) or ER+ (> 6.5). Recurrence Score results from the 21-gene assay were reported on a continuous scale from 0 to 100. A sub-analysis examined the association between ER expression (Allred score 2-7) and response to a 14-day pre-surgery pulse with an aromatase inhibitor. A separate sub-analysis examined the association between ER expression and human epidermal growth factor receptor 2 (HER2) expression. RESULTS: Tumor specimens from 192 patients (aged 25-92 years) were included in the pooled analysis. Correlation between IHC- and RT-PCR-measured ER was strong for IHC-defined ER-negative and ER+ samples (r = 0.646 [95% CI 0.553-0.720]). There was 100% concordance for ER+ tumors; however, 56% of the ER-low+ tumors were negative by RT-PCR. Allred score correlated better with ER status measured by RT-PCR at pre-treatment (r = 0.83) than at post-treatment (r = 0.76). The majority (77%) of ER-negative and ER-low+ tumors were HER2-negative. CONCLUSIONS: RT-PCR provided a more accurate assessment of ER expression in patients with ER-low+ tumors, and data support dual testing for patients with ER-low+ status to ensure appropriate treatment planning as it pertains to endocrine therapy. FUNDING: Genomic Health, Inc.

21-Gene assay as predictor of chemotherapy benefit in HER2-negative breast cancer.

The NSABP B-20 prospective-retrospective study of the 21-gene Oncotype DX Breast Cancer Recurrence Score® test predicted benefit from addition of chemotherapy to tamoxifen in node-negative, estrogen-receptor positive breast cancer when recurrence score (RS) was ≥31. HER2 is a component of the RS algorithm with a positive coefficient and contributes to higher RS values. Accrual to B-20 occurred prior to routine testing for HER2, so questions have arisen regarding assay performance if HER2-positive patients were identified and excluded. We report an exploratory reanalysis of the B-20, 21-gene study following exclusion of such patients. Patients were considered HER2 positive if quantitative RT-PCR for HER2 was ≥11.5 units, and excluded from re-analyses performed using the original cutoffs: <18, 18-30, ≥31, and the TAILORx cutoffs: <11, 11-25, >25. The endpoint remained distant recurrence-free interval (DRFI) as in the original study. Distribution was estimated via the Kaplan-Meier method and compared via log-rank test. Multivariate Cox proportional hazards models estimated chemotherapy benefit in each group. In the RS < 18 and 18-30 groups, 1.7 and 6.7% were HER2 positive. In the RS ≥ 31 group, 41% were HER2 positive. Exclusion resulted in fewer events, with loss of significance for benefit from chemotherapy in the overall HER2-negative cohort (log-rank P = 0.06), but substantial benefit from chemotherapy remained in the RS ≥ 31 cohort (HR = 0.18; 95% CI: 0.07-0.47) and the RS > 25 cohort (HR = 0.28; 95% CI: 0.12-0.64). No benefit from chemotherapy was evident in the other RS groups. Following exclusion of HER2-positive patients based on RT-PCR expression, substantial benefit of chemotherapy remained for RS ≥ 31 as originally employed, and with RS > 25 employed in TAILORx.

Erratum: Author Correction: Breast-cancer-specific mortality in patients treated based on the 21-gene assay: a SEER population-based study.

[This corrects the article DOI: 10.1038/npjbcancer.2016.17.].

The 21-gene Recurrence Score® assay predicts distant recurrence in lymph node-positive, hormone receptor-positive, breast cancer patients treated with adjuvant sequential epirubicin- and docetaxel-based or epirubicin-based chemotherapy (PACS-01 trial).

BACKGROUND: The 21-gene Recurrence Score (RS) result predicts outcome and chemotherapy benefit in node-negative and node-positive (N+), estrogen receptor-positive (ER+) patients treated with endocrine therapy. The purpose of this study was to evaluate the prognostic impact of RS results in N+, hormone receptor-positive (HR+) patients treated with adjuvant chemotherapy (6 cycles of FEC100 vs. 3 cycles of FEC100 followed by 3 cycles of docetaxel 100 mg/m2) plus endocrine therapy (ET) in the PACS-01 trial (J Clin Oncol 2006;24:5664-5671). METHODS: The current study included 530 HR+/N+ patients from the PACS-01 parent trial for whom specimens were available. The primary objective was to evaluate the relationship between the RS result and distant recurrence (DR). RESULTS: There were 209 (39.4%) patients with low RS (< 18), 159 (30%) with intermediate RS (18-30) and 162 (30.6%) with high RS (≥ 31). The continuous RS result was associated with DR (hazard ratio = 4.14; 95% confidence interval: 2.67-6.43; p <  0.001), adjusting for treatment. In multivariable analysis, the RS result remained a significant predictor of DR (p <  0.001) after adjustment for number of positive nodes, tumor size, tumor grade, Ki-67 (immunohistochemical status), and chemotherapy regimen. There was no statistically significant interaction between RS result and treatment in predicting DR (p = 0.79). CONCLUSIONS: After adjustment for clinical covariates, the 21-gene RS result is a significant prognostic factor in N+/HR+ patients receiving adjuvant chemoendocrine therapy. TRIAL REGISTRATION: Not applicable.

Comparison of the Performance of 6 Prognostic Signatures for Estrogen Receptor-Positive Breast Cancer: A Secondary Analysis of a Randomized Clinical Trial.

IMPORTANCE: Multiple molecular signatures are available for managing estrogen receptor (ER)-positive breast cancer but with little direct comparative information to guide the patient's choice. OBJECTIVE: To conduct a within-patient comparison of the prognostic value of 6 multigene signatures in women with early ER-positive breast cancer who received endocrine therapy for 5 years. DESIGN, SETTING, AND PARTICIPANTS: This retrospective biomarker analysis included 774 postmenopausal women with ER-positive ERBB2 (formerly HER2)-negative breast cancer. This analysis was performed as a preplanned secondary study of data from the Anastrozole or Tamoxifen Alone or Combined randomized clinical trial comparing 5-year treatment with anastrozole vs tamoxifen with 10-year follow-up data. The signatures included the Oncotype Dx recurrence score, PAM50-based Prosigna risk of recurrence (ROR), Breast Cancer Index (BCI), EndoPredict (EPclin), Clinical Treatment Score, and 4-marker immunohistochemical score. Data were collected from January 2009, through April 2015. MAIN OUTCOMES AND MEASURES: The primary objective was to compare the prognostic value of these signatures in addition to the Clinical Treatment Score (nodal status, tumor size, grade, age, and endocrine treatment) for distant recurrence for 0 to 10 years and 5 to 10 years after diagnosis. Likelihood ratio (LR) statistics were used with the χ2 test and C indexes to assess the prognostic value of each signature. RESULTS: In this study of 774 postmenopausal women with ER-positive, ERBB2-negative disease (mean [SD] age, 64.1 [8.1] years), 591 (mean [SD] age, 63.4 [7.9] years) had node-negative disease. The signatures providing the most prognostic information were the ROR (hazard ratio [HR], 2.56; 95% CI, 1.96-3.35), followed by the BCI (HR, 2.46; 95% CI, 1.88-3.23) and EPclin (HR, 2.14; 95% CI, 1.71-2.68). Each provided significantly more information than the Clinical Treatment Score (HR, 1.99; 95% CI, 1.58-2.50), the recurrence score (HR, 1.69; 95% CI, 1.40-2.03), and the 4-marker immunohistochemical score (HR, 1.95; 95% CI, 1.55-2.45). Substantially less information was provided by all 6 molecular tests for the 183 patients with 1 to 3 positive nodes, but the BCI (ΔLR χ2 = 9.2) and EPclin (ΔLR χ2 = 7.4) provided more additional prognostic information than the other signatures. CONCLUSIONS AND RELEVANCE: For women with node-negative disease, the ROR, BCI, and EPclin were significantly more prognostic for overall and late distant recurrence. For women with 1 to 3 positive nodes, limited independent information was available from any test. These data might help oncologists and patients to choose the most appropriate test when considering chemotherapy use and/or extended endocrine therapy. TRIAL REGISTRATION: isrctn.com Identifier: ISRCTN18233230.

21-Gene Recurrence Score for prognosis and prediction of taxane benefit after adjuvant chemotherapy plus endocrine therapy: results from NSABP B-28/NRG Oncology.

BACKGROUND: The 21-gene recurrence score (RS) predicts outcome and benefit from adjuvant chemotherapy benefit in breast cancer patients treated with adjuvant endocrine therapy. In the NSABP B-28 study, we evaluated the 21-gene RS for its prognostic impact and its ability to predict benefit from paclitaxel (P) in node-positive, estrogen receptor-positive (ER+) breast cancer patients treated with adjuvant chemotherapy plus tamoxifen. METHODS: The B-28 trial compared doxorubicin/cyclophosphamide (AC) with AC followed by P in 3060 patients. Tamoxifen for 5 years was also given to patients > 50 years and those < 50 years with ER+ and/or progesterone receptor-positive (PR+) tumors. The present study includes 1065 ER-positive, tamoxifen-treated patients with RS assessment. Median follow-up time was 11.2 years. RESULTS: In univariate analyses, RS was a significant predictor of outcome. In multivariate analyses, RS remained a significant independent predictor of outcome beyond clinico-pathologic factors, age, and type of surgery (p < 0.001). In the study population (n = 1065), the disease-free survival (DFS) hazard ratio (HR) with adding P to AC was 0.87 (95% CI 0.72-1.05; p = 0.14). RS was not a significant predictor of P benefit: for DFS, HRs for adding P to AC in RS low, intermediate, and high subgroups were 1.01 (95% CI 0.69-1.47; p = 0.99), 0.84 (95% CI 0.62-1.14; p = 0.26), and 0.81 (95% CI 0.60-1.10; p = 0.21), respectively (interaction p = 0.64). Similar findings were observed for the other study endpoints. CONCLUSIONS: RS maintains significant prognostic impact in ER-positive, node-positive patients treated with adjuvant chemotherapy plus tamoxifen. However, RS did not significantly predict benefit from adding paclitaxel to AC chemotherapy. (Trial Registration: PDQ: NSABP-B-28).