This secondary analysis of a randomized clinical trial evaluates the prognostic value of 6 multigene signatures in women with early estrogen receptor–positive breast cancer who have received 5 years of endocrine therapy.

Triple-negative breast cancers (TNBCs) have poor prognosis and lack targeted therapies. Interrogation of genomic datasets identified increased PIM1 copy number-driven expression in TNBC. RNA interference in breast cancer and non-malignant mammary epithelial cell models revealed a PIM1 dependency in TNBC cells for proliferation and apoptotic protection, absent in non-malignant cells. PIM1 knockdown reduced BCL2 expression, and dynamic BH3 profiling analysis revealed that PIM1 prevents mitochondrial-mediated apoptosis in TNBC cell lines. PIM1 expression associates with several MYC-transcriptional signatures and promotes cell population growth through regulation of c-MYC and transcription of MYC-targets, including MCL1. The pan-PIM kinase inhibitor AZD1208 inhibited growth and sensitized TNBC cell lines, xenografts and patient-derived xenografts to standard of care chemotherapy. We identify PIM1 as a malignant cell-selective target in TNBC, illustrating relationships with MYC activation, regulation of anti-apoptotic BCL2 and MCL1, established TNBC oncogenic proteins SHP2 and EPHA2 and cell cycle inhibitor p27. Finally we identify a potential use of PIM1 inhibitors to abrogate TNBC's high threshold to TNBC standard of care chemotherapy induced apoptotic cell death.

Background: Estimating distant recurrence (DR) risk among women with estrogen receptor–positive (ER+), human epidermal growth factor receptor 2 (HER2)–negative early breast cancer helps decisions on using adjuvant chemotherapy. The 21-gene Oncotype DX recurrence score (RS) is widely used for this. EndoPredict (EPclin) is an alternative test combining prognostic information from an eight-gene signature (EP score) with tumor size and nodal status. We compared the prognostic information provided by RS and EPclin for 10-year DR risk.Methods: We used likelihood ratio χ² and Kaplan-Meier survival analyses to compare prognostic information provided by EP, EPclin, RS, and the clinical treatment score (CTS) of clinicopathologic parameters in 928 patients with ER+ disease treated with five years’ anastrozole or tamoxifen. Comparisons were made for early (0-5 years) and late (5-10 years) DR according to nodal status. All statistical tests were two-sided.Results: In the overall population, EP and EPclin provided substantially more prognostic information than RS (LRχ2: EP = 49.3; LRχ2: EPclin = 139.3; LRχ2: RS = 29.1), with greater differences in late DR and in node-positive patients. EP and EPclin remained statistically significantly prognostic when adjusted for RS (ΔLRχ2: EP+RS vs RS = 20.2; ΔLRχ2: EPclin+RS vs RS = 113.8). Using predefined cut-offs, EPclin and RS identified 58.8% and 61.7% patients as low risk, with hazard ratios for non-low vs low risk of 5.99 (95% confidence interval [CI] = 3.94 to 9.11) and 2.73 (95% CI = 1.91 to 3.89), respectively.Conclusions: EP and EPclin were highly prognostic for DR in endocrine-treated patients with ER+, HER2-negative disease. EPclin provided more prognostic information than RS. This was partly but not entirely because of EPclin integrating molecular data with nodal status and tumor size.

Purpose: Phosphoinositide 3-kinase (PI3K) signaling is well established as important in cancer. To date most studies have been focused on the PI3K/p110α isoform, which has been found to be mutated in several different cancers. The aim of our study was to determine which specific PI3K isoforms are involved in pancreatic ductal adenocarcinoma (PDAC) and investigate the effects of these isoforms on proliferation, survival, and induction of Akt activation in pancreatic cancer cells.Experimental Design: The expression of all PI3K isoforms and downstream targets was analyzed by immunohistochemistry in human pancreatic cancer tissue and normal counterparts. Isoform selective inhibitors and short interfering RNA (siRNA) were employed to investigate the effects of the different PI3Ks on proliferation, survival, and intracellular signaling in PDAC cell lines.Results: Immunohistochemical screening revealed high specific expression of the PI3K/p110γ isoform. Scoring indicated that 72% of the PDAC tissue stained positive for PI3K/p110γ, whereas no stain was detected in normal pancreatic ducts. Proliferation analyses after selective inhibition and siRNA downregulation of PI3K/p110γ showed that PI3K/p110γ, but not other PI3K isoforms, was required for cell proliferation. Overexpression of PI3K/p110γ indeed increased cell numbers and mediated activation of Akt in PDAC cell lines. Moreover, PI3K/p110γ was required for Akt activation via lysophosphatidic acid receptors.Conclusions: These data represent the first identification of a tumor-specific accumulation of the PI3K isoform p110γ in human cancer. Further, our results signify a critical role for PI3K/p110γ in pancreatic cancer, and we hypothesize that PI3K/p110γ overexpression is a key event in the disease progression. Clin Cancer Res; 16(20); 4928-37. ©2010 AACR.

PURPOSE The Onco type 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.

Purpose EndoPredict (EPclin) is a prognostic test validated to inform decisions on adjuvant chemotherapy to endocrine therapy alone for patients with oestrogen receptor-positive, HER2-negative breast cancer. Here, we determine the performance of EPclin for estimating 10-year distant recurrence-free interval (DRFI) rates for those who received adjuvant endocrine therapy (ET) alone compared to those with chemotherapy plus endocrine therapy (ET + C). Methods A total of 3746 women were included in this joint analysis. 2630 patients received 5 years of ET alone (ABCSG-6/8, TransATAC) and 1116 patients received ET + C (GEICAM 2003-02/9906). The primary objective was to evaluate the ability of EPclin to provide an estimate of the 10-year DR rate as a continuous function of EPclin separately for ET alone and ET + C. Cox proportional hazard models were used for these analyses. Results EPclin was highly prognostic for DR in women who received ET alone (HR 2.79 (2.49–3.13), P < 0.0001) as well as in those who received ET + C (HR 2.27 (1.99–2.59), P < 0.0001). Women who received ET + C had significantly smaller increases in 10-year DR rates with the increasing EPclin score than those receiving ET alone (EPclin = 5; 12% ET + C vs. 20% ET alone). We observed a significant positive interaction between EPclin and treatment groups ( P - interaction = 0.022). Conclusions In this comparative non-randomised analysis, the rate of increase in DR with EPclin score was significantly reduced in women who received ET + C versus ET alone. Our indirect comparisons suggest that a high EPclin score can predict chemotherapy benefit in women with ER-positive, HER2-negative disease. Electronic supplementary material The online version of this article (10.1007/s10549-019-05226-8) contains supplementary material, which is available to authorized users.

SummaryThe scattering response of epithelial cells to activation of the Met receptor tyrosine kinase represents one facet of an “invasive growth” program [1, 2]. It is a complex event that incorporates loss of cell-cell adhesion, morphological changes, and cell motility. Ubiquitination is a reversible posttranslational modification that may target proteins for degradation or coordinate signal transduction pathways [3, 4]. There are ∼79 active deubiquitinating enzymes (DUBs) predicted in the human genome [5, 6]. Here, via a small interfering RNA (siRNA) library approach, we have identified 12 DUBs that are necessary for aspects of the hepatocyte growth factor (HGF)-dependent scattering response of A549 cells. Different phenotypes are evident that range from full loss of scattering, similar to receptor knockdown (e.g., USP30, USP33, USP47), to loss of cell-cell contacts even in the absence of HGF but defective motility (e.g., USP3, ATXN3L). The knockdowns do not incur defective receptor, phosphatidylinositol 3-kinase, or MAP kinase activation. Our data suggest widespread involvement of the ubiquitin system at multiple stages of the Met activation response, implying significant crosstalk with phosphorylation-based transduction pathways. Development of small-molecule inhibitors of particular DUBs may offer a therapeutic approach to contain metastasis.

Purpose: To identify the individual genes or gene modules that lead to the OncoptypeDx 21-gene recurrence score's reduced performance after 5 years and thereby identify indices of residual risk that may guide selection of patients for extended adjuvant therapy.Experimental Design: We conducted a retrospective assessment of the relationship between (i) the individual genes and gene modules of the Recurrence Score and (ii) early (0-5 years) and late (5-10 years) recurrence rates in 1,125 postmenopausal patients with primary estrogen receptor-positive breast cancer treated with anastrozole or tamoxifen in the Arimidex, Tamoxifen, Alone or Combined (ATAC) randomized clinical trial.Results: In the HER2-negative population (n ¼ 1,009), estimates of recurrence risk were similar between years 0-5 and 5-10 for proliferation and invasion modules but markedly different for the estrogen module and genes within it (all split at the median): for low estrogen module, annual recurrence rates were similar across the two time windows (2.06% vs. 2.46%, respectively); for high estrogen module, annual rates were 1.14% versus 2.72%, respectively (P interaction ¼ 0.004). Estrogen receptor transcript levels showed inverse prediction across the time windows: HR, 0.88 (0.73-1.07) and 1.19 (0.99-1.43), respectively (P interaction ¼ 0.03). Similar time-, module-, and estrogen-dependent relationships were seen for distant recurrence.Conclusions: Patients with tumors with high estrogen receptor transcript levels benefit most from 5 years' endocrine therapy but show increased recurrence rates after 5 years and may benefit from extended therapy. Improved prognostic profiles may be created by considering period of treatment and follow-up time.