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Background: In the era of targeted therapies, chemotherapy (CT) is still a valuable treatment option for patients (pts) with HER2 negative metastatic breast cancer (MBC). The identification of predictive and prognostic markers might improve treatment response and survival. Here we investigated the role of tumor subtypes, circulating tumor cells (CTCs), and mutations in genes or pathways of interest, in predicting response to CT and prognosis of HER2 negative MBC pts within AARES trial. Methods: AARES is an open label multicentric randomised phase 2 trial comparing a DNA-damaging (arm A: cisplatin 25 mg/m2/day, day 1-3 + cyclophosphamide 600 mg/m2 day 1) versus (vs) a non-DNA-damaging (arm B: capecitabine 1000 mg/m2 bid/day, day 1-14 + vinorelbine 60 mg/m2/day, day 1,8) CT regimen in pts with HER2 negative MBC. Archival tumor tissue samples and blood samples were collected at baseline. The Cell SearchTM system was used for CTCs isolation and enumeration and a cut-off of 5 CTC/7,5 ml was used. Tumor subtypes were based on locally assessed hormone receptors and HER2 status. The luminal-like subtype (Lum) was defined by ER or PR positivity, while the triple negative subtype (TN) by ER, PR, and HER2 negativity. Tissue samples were used for DNA extraction and next generation sequencing analysis using a target enrichment panel of 170 genes (TruSight 170, Illumina). Primary endpoint was objective response rate (ORR) assessed for the two treatment arms. Progression free survival (PFS) and overall survival (OS) were evaluated as secondary endpoints, and estimated with the Kaplan-Meier method. ORR, PFS and OS were correlated with CTC counts, tumor subtypes, and mutational status of genes/pathway of interest (PIK3CA, TP53, BRCA1, BRCA2, PI3K/AKT pathway (PI3K/AKT)). Results: AARES enrolled 102 pts from 2011 to 2016 across 9 Italian centers. Of these, 77 pts were evaluable for ORR. Median follow up was 32 months. Overall, median age was 57. The majority of pts had Lum tumors (73%), while 27% had TN. 86% of pts had visceral metastases, and 52% had 3 or more metastatic sites. 48% of pts received the study treatment as 1st line, 32% as 2nd line, and 19% as 3rd line. Out of the 77 pts with ORR data, 41 (53%) were CTC- and 36 (47%) CTC+. A larger proportion of pts with Lum tumors were CTC+ (54% n=30), while TN tumors were mainly CTC- (71% n=15). 49 pts had adequate tumor tissue for sequencing analysis. TP53 mutations (mut) were found in 33% of pts, BRCA1/2 mut in 14%, PIK3CA mut in 35%, and PI3K/AKT mut in 47%. ORR was 24% and 37% in arm A vs arm B, respectively (p= 0.2), and no difference in ORR was observed by tumor subtypes (Lum vs TN), and CTCs count (+ vs -). PFS and OS were assessable on the whole population (n=102) and did not significantly differ by treatment arms. Median OS (mOS) was higher in Lum pts compared to TN (24 months (mo) vs 15.4 mo, p=0.048) while median PFS (mPFS) did not differ according to tumor subtypes (p=0.28). Overall, CTCs count was not significantly associated with mPFS or OS however in pts with TN tumors and CTC-, mPFS and mOS were significantly longer (mPFS: 4.44 mo vs 2.56 mo in CTC- and CTC+ subgroups, respectively, p=0.00087 and mOS: 22.07 mo vs 3.57 mo in CTC- and CTC+ subgroups, respectively, p< 0.0001). On the other hand, no differences were observed in Lum pts categorized according to CTC status (p=0.83 for mPFS, p=0.54 for mOS). Finally, pts with PI3K/AKT mut had a significantly worse PFS compared to wild type in the overall population (p=0.0091) as well as in Lum (p=0.034) and TN pts (p=0.0052). Conclusions: CTCs and tumor subtypes were not predictive of response to CT regimens in HER2 negative MBC pts. A number of CTC >5 in TN MBC, and mutations in the PI3K/AKT pathway, identified a subgroup of pts with worse prognosis, potentially candidates for alternative treatments. Further studies are needed to confirm these results in a larger population. Citation Format: Emanuela Risi, Marta Pestrin, Chiara Biagioni, Dario Romagnoli, Ilenia Migliaccio, Francesca Galardi, Francesca De Luca, Matteo Benelli, Erica Moretti, Giuseppina Sanna, Luca Livraghi, Silvia Cappadona, Roberta Di Marsico, Domenico Amoroso, Angelo Martignetti, Angela S. Ribecco, Elena Rota Caremoli, Luigi Coltelli, Fabio Puglisi, Luca Malorni, Laura Biganzoli. Circulating tumor cells, immunohistochemical subtypes, and genes mutation as prognostic markers in HER2 negative metastatic breast cancer patients candidates to chemotherapy [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 P3-05-45.

Baseline characteristics of studied c-TREnd biomarker population - including all patients with at least one sample available for analysis at any timepoint(s), compared to the overall original TREnd cohort. Fisher exact test was used to calculate p-values. *Denotes patients included in intention-to-treat population of TREnd, despite breach of eligibility criteria (discovered post-enrolment).

Purpose:Thymidine kinase 1 (TK1) is downstream to the CDK4/6 pathway, and TK activity (TKa) measured in blood is a dynamic marker of outcome in patients with advanced breast cancer (ABC). This study explores TK1 as a biomarker of palbociclib response, both in vitro and in patients with ABC.Experimental Design:Modulation of TK1 levels and activity by palbociclib were studied in seven estrogen receptor–positive breast cancer cell lines: sensitive (PDS) and with palbociclib acquired resistance (PDR). TKa was assayed in plasma obtained at baseline (T0), after one cycle (T1), and at disease progression on palbociclib (T2) in patients enrolled in the “To Reverse ENDocrine Resistance” (TREnd) trial (n = 46).Results:Among E2F-dependent genes, TK1 was significantly downregulated after short-term palbociclib. Early TKa reduction by palbociclib occurred in PDS but not in PDR cells. In patients, median TKa (mTKa) at T0 was 75 DiviTum units per liter (Du/L), with baseline TKa not proving prognostic. At T1, mTKa decreased to 35 Du/L, with a minority of patients (n = 8) showing an increase—correlating with a worse outcome than those with decreased/stable TKa (n = 33; mPFS 3.0 vs 9.0 months; P = 0.002). At T2, mTKa was 251 Du/L; patients with TKa above the median had worse outcomes on post-study treatment compared with those with lower TKa (2.9 vs 8.7 months; P = 0.05).Conclusions:TK is a dynamic marker of resistance to palbociclib which may lead to early identification of patients in whom treatment escalation may be feasible. In addition, TKa may stratify prognosis in patients with acquired resistance to palbociclib.

The addition of supplemental food sources for natural enemies is a growing component of conservation and augmentative biological control. Supplemental foods can be used to retain natural enemies when prey are scarce and to promote survival, fecundity, longevity, and development of natural enemy populations, especially generalist natural enemies. Amblydromella caudiglans (Schuster) (Acari: Phytoseiidae) is one of the most abundant predatory mites found in commercial apple orchards in Washington, USA, and contributes to spider mite control. However, because its widespread presence in commercial apple orchards was only recently discovered, how supplementary food sources affect its performance is unknown. In laboratory studies, we evaluated the performance (fecundity, retention, prey consumption) of the generalist phytoseiid A. caudiglans on commercially available supplemental food sources, including factitious prey (Ephestia eggs and Artemia brine shrimp cysts), and pollens of apple, pear, and cattail. We determined that A. caudiglans will not consume Ephestia eggs and performs best on cattail and pear pollens. Combinations of food sources did not enhance the performance of this predator compared to the best performing single-sources. The presence of alternative food sources did not decrease A. caudiglans predation of twospotted spider mite nymphs, except for Artemia brine shrimp cysts, which had a substantial handling time. These results lay the groundwork for identifying a way to promote and retain this natural enemy in tree fruit cropping systems through the use of food resource applications or floral plantings.

Introduction: Breast cancer (BC) is a heterogeneous disease characterized by different molecular "intrinsic" subtypes (PAM50) with distinct prognostic and therapeutic implications. DNA methylation (DNAm) is one of the most studied epigenetic mechanisms involved in BC tumorigenesis and progression. Recently, many studies have demonstrated the capability of DNAm-based assays in detecting and monitoring the disease in liquid biopsy samples from patients with BC, thus confirming promising role of DNAm-based biomarkers in the context of precision oncology. Here we present a novel computational method that exploits a minimal set of informative, clonal DNAm sites to estimate tumor content and molecular subtype in BC tissue samples with potential application for liquid biopsy analyses. Methods: DNAm data of TCGA-BRCA comprising 737 tumor samples, including 135 Basal-like (Basal), 46 HER2-enriched (HER2e) and 556 Luminal A/B (LumA/B) samples, and 96 normal tissue samples were downloaded from GDC Data Portal. To estimate tumor purity, we selected DNAm sites based on the following characteristics: (1) AUC>0.8 or AUC350 samples, high accuracy was obtained for LumA/B and Basal samples while HER2e samples showed lower F1-score due to a drop of recall, though obtaining precision >80% in both datasets. Based on these results, a DNAm-based targeted-seq assay has been designed and will be applied to a series of plasma samples from patients with metastatic BC collected within our center. First results of the assay will be presented during the meeting. Citation Format: Dario Romagnoli, Francesca Galardi, Francesca De Luca, Chiara Biagioni, Erica Moretti, Laura Biganzoli, Ilenia Migliaccio, Luca Malorni, Matteo Benelli. A minimal DNA-methylation signature to estimate tumor content and molecular subtype in breast cancer tissue samples with potential application to liquid biopsy [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P3-05-02.

We previously showed that metabolomics predicts relapse in early breast cancer (eBC) patients, unselected by age. This study aims to identify a "metabolic signature" that differentiates eBC from advanced breast cancer (aBC) patients, and to investigate its potential prognostic role in an elderly population.Serum samples from elderly breast cancer (BC) patients enrolled in 3 onco-geriatric trials, were retrospectively analyzed via proton nuclear magnetic resonance (1H NMR) spectroscopy. Three nuclear magnetic resonance (NMR) spectra were acquired for each serum sample: NOESY1D, CPMG, Diffusion-edited. Random Forest (RF) models to predict BC relapse were built on NMR spectra, and resulting RF risk scores were evaluated by Kaplan-Meier curves.Serum samples from 140 eBC patients and 27 aBC were retrieved. In the eBC cohort, median age was 76 years; 77% of patients had luminal, 10% HER2-positive and 13% triple negative (TN) BC. Forty-two percent of patients had tumors2 cm, 43% had positive axillary nodes. Using NOESY1D spectra, the RF classifier discriminated free-from-recurrence eBC from aBC with sensitivity, specificity and accuracy of 81%, 67% and 70% respectively. We tested the NOESY1D spectra of each eBC patient on the RF models already calculated. We found that patients classified as "high risk" had higher risk of disease recurrence (hazard ratio (HR) 3.42, 95% confidence interval (CI) 1.58-7.37) than patients at low-risk.This analysis suggests that a "metabolic signature", identified employing NMR fingerprinting, is able to predict the risk of disease recurrence in elderly patients with eBC independently from standard clinicopathological features.

Background: Thymidine kinase-1 is a cell proliferation marker downstream of the CDK4/6 pathway, whose activity can be measured in serum to reflect tumor proliferation. The CDK4/6 inhibitor palbociclib (P) is approved for the treatment of patients (pts) with hormone receptor positive metastatic breast cancer (MBC) in first or second line endocrine-based treatment settings. Approximately 10-15% of pts exhibit de novo resistance to P, with circulating levels of thymidine kinase activity (TKa) previously shown as a potential marker of early treatment resistance. Therapeutic strategies to address primary resistance to P are currently lacking. Little is known of the clinical efficacy of alternative dosing schedules of P, and its effect on TKa. Here we report serum TKa measured at different timepoints from samples collected within the MA38 (NCT02630693) study. Methods: MA38 is an open label randomised Phase 2 trial comparing two different schedules of P plus second-line ET in pts with ER-positive, HER2-negative MBC. Pts were assigned to receive physician’s choice ET plus either standard P dosing (125mg daily for 21 days on a 28-day cycle), or 100mg daily continuously. Serum samples were collected at baseline (BL; n=135), at 12 weeks (W12; n=122) and 24 weeks (W24; n=95). TKa was measured with DiviTum®, a refined ELISA-based assay (lower limit of detection [LLOD] = 100 DuA). Kaplan-Meier method estimated BL, W12 and W24 (95% CI) median PFS (mPFS; from randomization until progression by RECIST criteria or death) and overall survival (OS; from randomization until death from any cause) in groups of patients defined by dichotomizing TKa as “high” or “low” at the median. Results: MA38 enrolled 180 pts from December 2015 and February 2017 across Canada. Median follow up was 19 months. Overall, the median age was 60, and 90% of pts were post-menopausal. All pts had estrogen receptor-positive disease, and 64% had visceral metastases. On study, 56% received fulvestrant with P, 34% aromatase inhibitor and 10% tamoxifen. TKa was successfully measured in 100% of samples. Median TKa (mTKa) at BL was 234 DuA (IQR 138.5 - 438). BL TKa was not associated with clinical or pathological characteristics. TKa was prognostic at BL with mPFS of 5.5 months (mo) in pts with high TKa vs 16.3 mo with low TKa (HR=2.43; 95% CI, 1.6-3.7; p< 0.001). Similar results were obtained employing other previously reported cut off values. At multivariate analysis, BL TKa was independent from other prognostic factors including age, ECOG status and presence of visceral metastases (adjusted HR= 2.34; 95%CI 1.5- 3.6; p < 0.001). In terms of OS, BL TKa was an independent prognostic factor (adjusted HR=2.0; 95% CI, 1.1-3.7; p=0.02). At 12 mo, OS rate was 68% in pts with high BL TKa vs 92% in low TKa. Both for PFS and OS, no interaction between BL TKa and study arm was observed. At W12 mTKa was 129.5 DuA (IQR 100 - 219.8) and below LLOD (IQR 100 - 180) at W24. At these timepoints, landmark analyses showed no significant difference in PFS according to TKa. However, at W12 high TKa was significantly associated with worse OS (HR 2.0; 95%CI 1.0- 4.0; p=0.03), with a similar trend at W24 (HR 2.5; 95%CI 0.9-6.4; p=0.06). Conclusions: Baseline TKa is a reliable prognostic marker of both PFS and OS in pts treated with P and ET, further substantiating previous data. Monitoring TKa during treatment may provide important clinical information. A significant relationship between TKa and assigned treatment arm was not observed, suggesting TKa is not influenced by P treatment dose or intensity. These data confirm the role of baseline TKa as a new marker for patient stratification, and supports further investigation for the assessment of the clinical utility of TKa as a monitoring biomarker in the advanced setting. Citation Format: Amelia McCartney, Chiara Biagioni, Bingshu Chen, Lois Shepherd, Karen Gelmon, Anil A. Joy, Wendy Parulekar, Mattias Bergqvist, Ilenia Migliaccio, Angela Leo, Matteo Benelli, Emanuela Risi, Erica Moretti, Luca Livraghi, Laura Biganzoli, Luca Malorni. Serum thymidine kinase activity as a prognostic marker in women with metastatic breast cancer treated with two different schedules of palbociclib plus second-line endocrine therapy within the CCTG MA38 trial [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 P5-02-27.

Background: We have previously identified two potentially predictive signatures of palbociclib resistance: the RBsig, composed of E2F1/E2F2 dependent genes, which is correlated with genetic loss of RB1, and the ratio between the gene expression levels of CCNE1 to RB1 (CCNE1/RB1). Both signatures have been previously tested in vitro and in neoadjuvant studies with palbociclib. The present analysis aims to explore the prognostic and predictive role of RBsig and CCNE1/RB1 in the pivotal phase III randomized trials PALOMA-2 and PALOMA-3. Materials and methods: Gene expression data from the PALOMA-2 and PALOMA-3 datasets were generated using the HTG EdgeSeq Oncology BM Panel, as previously described. Of the 87 genes composing RBsig, 46 were available within the EdgeSeq dataset and were used for the analyses; CCNE1 and RB1 were both available. RBsig was calculated as the mean of the Z-score scaled gene expression (log) of the 46 genes; CCNE1/RB1 was computed as the log ratio between the mRNA expression of CCNE1 and RB1. High and low values of RBsig and CCNE1/RB1 were defined based on the third quartile (Q3) as cutoff or as continuous variables. The prognostic/predictive effect of the signatures in terms of PFS was tested using Cox proportional hazard models and the Wald test. Results: The 46-genes RBsig versus the original signature showed excellent correlation in the METABRIC dataset (R=0.99), confirming its reliability as a surrogate of the original RBsig using EdgeSeq data. In both PALOMA-2 and PALOMA-3, RBsig high was significantly associated with a worse outcome compared to RBsig low in the palbociclib arm but not in the control arm [PALOMA-2: HR 1.4 (95% CI 1.0, 2.0) p=0.029 for palbociclib arm; HR 1.1 (95% CI 0.7, 1.6) p=0.71 for control arm. PALOMA-3: HR 1.7 (95% CI 1.1, 2.6) p=0.01 for palbociclib arm; HR 1.2 (95% CI 0.7, 1.9) p= 0.49 for control arm]. However, in both studies RBsig was not predictive of palbociclib resistance both when considered as a continuous variable and when dichotomized at Q3. Similarly to RBsig, in PALOMA- 3 patients with CCNE1/RB1 high tumors treated in the palbociclib arm showed a significantly worse outcome compared to those with CCNE1/RB1 low but this effect was not observed in those treated in the control arm [HR 1.6 (95% CI 1.1- 2.5) p= 0.03 for palbociclib arm; HR 1.2 (95% CI 0.7, 1.9) p=0.5 for control arm]. In addition, CCNE1/RB1 as a continuous variable was predictive of palbociclib benefit in PALOMA-3 (interaction p= 0.047). These effects were not observed in PALOMA-2. Conclusions: RBsig is a prognostic biomarker in patients treated with palbociclib, suggesting it may help in patients’ risk stratification. CCNE1/RB1 is predictive of palbociclib benefit in PALOMA-3, but not in PALOMA-2 probably due to the different patient populations and characteristics. Further studies of these biomarkers in patients treated with CDK4/6 inhibitors in the metastatic as well in the adjuvant setting are warranted. Citation Format: Luca Malorni, Matteo Benelli, Yuan Liu, Shibing Deng, Zhe Zhang, Cristina Guarducci, Angela Leo, Agostina Nardone, Francesca Galardi, Emanuela Risi, Erica Moretti, Dario Romagnoli, Chiara Biagioni, Marta Paoli, Laura Biganzoli, Ilenia Migliaccio. Prognostic and predictive role of RBsig and CCNE1/RB1 gene-expression signatures in patients with advanced breast cancer treated with palbociclib in combination with endocrine therapy in the PALOMA-2 and 3 trials [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 P5-02-17.

Precision oncology is an emerging approach in cancer care. It aims at selecting the optimal therapy for the right patient by considering each patient’s unique disease and individual health status. In the last years, it has become evident that breast cancer is an extremely heterogeneous disease, and therefore, patients need to be appropriately stratified to maximize survival and quality of life. Gene-expression tools have already positively assisted clinical decision making by estimating the risk of recurrence and the potential benefit from adjuvant chemotherapy. However, these approaches need refinement to further reduce the proportion of patients potentially exposed to unnecessary chemotherapy. Nuclear magnetic resonance (NMR) metabolomics has demonstrated to be an optimal approach for cancer research and has provided significant results in BC, in particular for prognostic and stratification purposes. In this review, we give an update on the status of NMR-based metabolomic studies for the biochemical characterization and stratification of breast cancer patients using different biospecimens (breast tissue, blood serum/plasma, and urine).

Liquid biopsy based on cell-free DNA (cfDNA) enables non-invasive dynamic assessment of disease status in patients with cancer, both in the early and advanced settings. The analysis of DNA-methylation (DNAm) from cfDNA samples holds great promise due to the intrinsic characteristics of DNAm being more prevalent, pervasive, and cell- and tumor-type specific than genomics, for which established cfDNA assays already exist. Herein, we report on recent advances on experimental strategies for the analysis of DNAm in cfDNA samples. We describe the main steps of DNAm-based analysis workflows, including pre-analytics of cfDNA samples, DNA treatment, assays for DNAm evaluation, and methods for data analysis. We report on protocols, biomolecular techniques, and computational strategies enabling DNAm evaluation in the context of cfDNA analysis, along with practical considerations on input sample requirements and costs. We provide an overview on existing studies exploiting cell-free DNAm biomarkers for the detection and monitoring of cancer in early and advanced settings, for the evaluation of drug resistance, and for the identification of the cell-of-origin of tumors. Finally, we report on DNAm-based tests approved for clinical use and summarize their performance in the context of liquid biopsy.

Background Metronomic chemotherapy can induce disease control in patients with metastatic breast cancer (MBC) and has better safety profiles than conventional chemotherapy. Evidence suggests that cytotoxics can be anti-angiogenic in pre-clinical models and may have synergistic effects when combined with anti-vascular endothelial growth factor therapies. Patients and Methods Patients pretreated with ≥ 1 prior line of therapy for MBC received oral cyclophosphamide 50 mg daily in combination with oral vinorelbine at escalating doses of 20 mg (V20), 30 mg (V30), and 40 mg (V40) 3 times per week, and intravenous bevacizumab 15 mg/kg every 3 weeks. Patients with human epidermal growth factor receptor 2-positive disease were given the same regimen plus standard trastuzumab. Doses were escalated when 3 patients completed 3 treatment cycles of V20 and V30, without experiencing dose-limiting toxicities. The recommended dose was then tested in a further 6 patients. Circulating tumour cells and circulating endothelial cells (CEC) were measured in 30 mL of whole blood samples at baseline, after cycle 1, and at the disease progression. Results Fifteen patients were recruited from June 2013 to October 2015. The median age was 61 years (range, 29-72 years); 80% had estrogen receptor-positive and 33% had human epidermal growth factor receptor 2-positive disease. At least 67% had visceral metastases, and 80% had received ≥ 2 lines of prior treatment. No dose-limiting toxicities were observed at the 3 dose-levels, making V40 the recommended dose. Overall 8 (53%) patients developed grade 2 adverse events (arthralgia, n = 3 [20%]; asthenia, n = 2 [13%]; diarrhea, n = 2 [13%]; leukopenia, n = 2 [13%]). Bevacizumab was associated with grade 3 hypertension (n = 3 [20%]). Stable disease as best response was observed in 11 (73.3%) patients. The clinical benefit rate was 66.6% (10/15 patients). The median time to progression was 6.9 months. At baseline, CECs were more commonly detectable than circulating tumor cells; however, no statistical correlation was found between CEC kinetics and response. Conclusion A metronomic vinorelbine dose of 40 mg combined with cyclophosphamide and bevacizumab is a promising treatment regimen in pretreated patients with MBC.

Background. Partial breast irradiation (PBI) is a reasonable alternative for a whole breast irradiation (WBI) in selected early stage breast cancer patients. The 5-year analysis of the APBI IMRT Florence phase 3 trial showed no significant difference in terms of ipsilateral tumor recurrence (IBTR) and survival rates between the two arms. We present the 10-year follow up data. Methods. From March 2005 to June 2013, women aged more than 40 years affected by early BC, with a maximum pathological tumor size of 25 mm, were randomly assigned in a 1:1 ratio to receive either WBI using three-dimensional conformal radiotherapy (3DCRT) or accelerated PBI (APBI) using intensity-modulated radiotherapy (IMRT) technique. Patients in the APBI arm received a total dose of 30 Gy to the tumor bed in five daily fractions. The WBI arm received 50 Gy in 25 fractions, followed by a boost on the tumor bed of 10 Gy in five fractions. The primary end-point, IBTR rate is now assessed at 10 years in the ITT population, as well as secondary endpoints: overall survival (OS), breast cancer specific survival (BCSS), distant metastasis free survival (DMFS), contralateral breast cancer (CBC), and locoregional recurrences (LRR). This trial is registered with ClinicalTrials.gov, number NCT02104895. Results. Of the 520 patients, 260 were enrolled in the APBI-arm and 260 in the WBI-arm; median age was 62.8 years (63.6 in the APBI and 61.6 in the WBI arm, p=0.20). Both treatment-arms achieved a median 10-year follow-up (10.1 in APBI-arm and 10.4 in WBI-arm, p=0.39) and were comparable regarding age, tumor size, grade, tumor type, and adjuvant endocrine treatment. No significant difference in terms of IBTR rates between treatment arms was shown (log rank test p=0.58). In the group who received APBI, the 5-year IBTR was 1.96% (5 events; 95% CI: 0.3;3.7) and the 10-year IBTR was 3.74% (9 events; 95% CI: 1.5;6.3). In the group who received WBI, the 5-year IBTR rate was 1.2% (3 events; 95% CI: 0;2.5) and the 10-year IBTR was 2.5% (6 events; 95% CI: 0.7;4.5). HR for APBI patients compared with WBI patients was 1.33 (p=0.58; 95% CI: 0.49;3.56). There were no significant differences between treatment arms regarding survival outcomes. OS (log rank test p=0.33): APBI 95.4% (95% CI: 93.6;97.2) versus WBI 94.3% (95% CI: 92.3;96.3); HR for APBI patients 0.66 (p=0.33; 95% CI: 0.29;1.53). BCSS (log rank test p=0.55): APBI 98% (95% CI: 96.8;99.2) versus WBI 97.5% (95% CI: 96.2;98.8); HR for APBI patients 0.68 (P=0.55; 95% CI: 0.19;2.42). DMFS (log rank test p=0.45): APBI 97.4% (95% CI: 96.0;98.8) versus WBI 96.1% (95% CI: 94.4;97.8); HR for APBI patients 0.67 (p=0.45; 95% CI: 0.24;1.89). Cumulative incidence estimates of 10-year LRR were 3.9% (95% CI: 2.24;5.56) for APBI versus 3.0% (95% CI: 1.53;4.47) for WBI (log rank test p=0.59). CBC observed after APBI were 5 (1.9%) as compared to 9 (3.5%) after WBI (p=0.42). Conclusion. IBTR rate after 10 years in patients with early breast cancer who were treated with APBI using IMRT technique in 5 fractions is rare and not significantly different from patients treated with WBI. OS, BCSS, DMFS, and LRR control are also comparable. Thus, APBI should be considered a reasonable alternative for a WBI in early breast cancer patients. Citation Format: Icro Meattini, Calogero Saieva, Sara Lucidi, Monica lo Russo, Vieri Scotti, Isacco Desideri, Livia Marrazzo, Gabriele Simontacchi, Monica Mangoni, Carlotta Becherini, Luca Visani, Lisa Paoletti, Erica Moretti, Luca Triggiani, Marco Bernini, Lorenzo Orzalesi, Luis Sanchez, Jacopo Nori, Stefania Pallotta, Simonetta Bianchi, Lorenzo Livi. Accelerated partial breast or whole breast irradiation after breast conservation surgery for patients with early breast cancer: 10-year follow up results of the APBI IMRT Florence randomized phase 3 trial [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 GS4-06.

Background The activity of palbociclib as a single agent in advanced breast cancer has not been extensively studied, with the only available clinical data limited to heavily pretreated patients. Preclinical data suggests palbociclib may partially reverse endocrine resistance, though this hypothesis has not been evaluated in previous clinical studies. This phase II, open-label, multicenter study examined the activity of palbociclib monotherapy, as well as palbociclib given in combination with the same endocrine therapy (ET) that was received prior to disease progression, in postmenopausal women with moderately pretreated, estrogen receptor-positive, HER2 negative advanced breast cancer. Patients and methods Eligible women with advanced disease which had progressed on one or two prior ETs were randomized 1 : 1 to receive either palbociclib alone, or palbociclib in combination with the ET as previously received. Primary end point was clinical benefit rate (CBR); secondary end points included progression-free survival (PFS). Results Between October 2012 and July 2016, a total of 115 patients were randomized. The CBR was 54% [95% confidence interval (CI): 41.5–63.7] for combination therapy, and 60% (95% CI: 47.8–72.9) for monotherapy. Median PFS was 10.8 months (95% CI: 5.6–12.7) for combination therapy, and 6.5 months (95% CI: 5.4–8.5) for monotherapy [hazard ratio (HR) 0.69; 95% CI: 0.4–1.1, exploratory P-value = 0.12]. Exploratory analyses revealed the PFS advantage for combination therapy was seen in the subgroup of patients who received prior ET for >6 months (HR 0.53; 95% CI: 0.3–0.9, exploratory P-value = 0.02), but not in those who received prior ET for ≤6 months. Conclusion Palbociclib has clinical activity as a single agent in women with moderately pretreated, oestrogen receptor-positive, HER2-negative advanced breast cancer. Palbociclib may have potential to reverse endocrine resistance in patients with a history of previous durable response to ET. Clinical trial information NCT02549430

Patients with luminal early breast cancer are at risk of relapse, even after five years of adjuvant endocrine therapy. To date, no biomarkers have been clinically validated to identify those patients at risk of late recurrence, who might benefit from extended adjuvant endocrine therapy. In recent decades, multiple clinical trials have tested the role of extending adjuvant endocrine therapies in patients with luminal disease. However, the data currently at our disposal are conflicting. This article reviews all the major trials concerning extended adjuvant endocrine regimens, and formulates some general conclusions and hypotheses of future study.

Background The combination of platinum with 5-fluorouracil has scarcely been studied in metastatic breast cancer. As this combination does not lead to significant hepatic metabolism, in some clinical situations it may prove useful, especially in cases with liver dysfunction and an urgent clinical need for rapid tumor shrinkage. A retrospective study was conducted to evaluate the efficacy and safety of the combination of cisplatin and 5-fluorouracil in patients with metastatic breast cancer with significant alterations of biochemistry. Patients and methods A total of 109 patients with metastatic breast cancer and liver dysfunction were treated; time-to-progression, overall survival and trends in liver function were evaluated. Results The median time-to-progression was 3.4 months, and median overall survival was 7.8 months. About 50% of patients obtained a complete, partial or stable biochemical response and 24 patients were subsequently able to receive additional therapies. Conclusion Our results show that this therapeutic doublet represents a clinically effective, safe and well-tolerated treatment option for patients with metastatic breast cancer and liver dysfunction.

Until recently, the mainstay of treatment in the majority of hormone receptor (HR)-positive, human epidermal growth factor 2 receptor (HER2)-negative advanced breast cancer (ABC) has consisted of single-agent endocrine therapy (ET). However, as understanding of endocrine resistance has grown, newer targeted agents have come to the fore. Inhibition of cyclin-dependent kinase complexes 4 and 6 (CDK4/6) combined with ET has shown significant activity in HR+ HER2− ABC, with impressive results in terms of progression-free survival (PFS) when compared with ET alone. This review summarizes the seminal findings pertaining to CDK4/6 inhibition in this population, specifically focusing on abemaciclib, contrasted with palbociclib and ribociclib. Potential directions for future studies are discussed, as a way of addressing outstanding issues such as establishing optimal treatment sequencing and agent combinations, appropriate patient selection to derive maximal benefits, predictive biomarkers and the employment of CDK4/6 inhibition beyond the ABC setting.

In the early years of the twentieth century, the great structural, social and cultural changes in American society included a growing number of immigrants arriving from the poorest regions of Europe. For the first time, the issues of immigration, assimilation and social integration became the most important problems facing American society. In the optimistic climate of the so-called progressive era, social reformers thought that these problems could be solved by the science of pedagogy, as applied to the educational needs of foreign immigrants. This essay centres on the pedagogical efforts of Italian-American educator Angelo Patri, who attempted to integrate Italian-American children into the fabric of American society through education. It starts by assessing Patri’s early writings, such as A Schoolmaster of the Great City, and his private and professional papers. In doing so, his work is situated in the debate on progressive education alongside pedagogue Maria Montessori, demonstrating his central role ...

Background and aim of the work: The treatment of cardiac arrest in an extra-hospital environment improves with the increase in the number of people able to establish an early Cardio-Pulmonary Reanimation (CPR). The main aim of the study was to assess the validity of the two-step method in case of prolonged CPR. Methods: A randomized comparison study was conducted in the University Nursing School of a Northern Italian town, during the 2015/16 academic year, among 60 students, to teach them CPR techniques, through two different teaching methods (4-step and the 2-step of CPR training). The effectiveness of the maneuvers performed on mannequins equipped with skill-meter was verified. Results: Our study did not highlight any significant difference between the two methods of CPR training. The comparison between the two methods regarding their efficacy in practical teaching of CPR, highlighted by this study, proved the validity of both the 4-minute continuous method (1st method) and the 30:2 method (2nd method). Conclusions: The results of the study showed no differences between the 2-step and the 4-step methods, in the effectiveness of cardiac massage. The correct execution of chest compressions during a CPR is the key to increase the patient’s chances of rescue. Research has shown that any interruption in the execution of chest compressions, leads to a progressive reduction of the effectiveness of cardiac massage, with negative consequences on the prognosis of the patient undergoing at CPR.

Background: The CDK4/6 inhibitor palbociclib (P) is approved for the treatment of luminal metastatic breast cancer (MBC) in combination with endocrine therapy (ET). It leads to reduced phosphorylation of the Rb protein resulting in a decrease in E2F activity and eventual cell cycle arrest. Thymidine kinase 1 is a well-known cancer proliferation marker downstream of the E2F pathway, whose activity can be measured in plasma samples as readout of tumor proliferation. Circulating thymidine kinase activity (TKa) is a prognostic marker in MBC patients (pts) treated with ET, both when measured at baseline and during treatment. TKa has been previously shown to decrease in pts treated with neoadjuvant P+ET for 15 days, which was attributed as pharmacodynamic change on P treatment. The predictive value of TKa changes during treatment with P, as well as the dynamics of TKa changes on P-containing treatments are not yet comprehensively defined. Here we investigate the role of plasma TKa measured at different timepoints in a cohort from the TREnd study (NCT02549430). Methods: TREnd was a randomized phase II trial that allocated 115 pts with moderately pre-treated luminal MBC to receive single-agent P or P plus the same ET agent that was received in the prior line of therapy. Plasma samples were collected at baseline (T0; n=45), at day 1 of cycle 2 (T1; n=45) and at disease progression (T2; n= 36) from 46 consenting pts. TKa was measured with DiviTum®, a refined ELISA-based assay. Patients were dichotomized as high/low at T0 based on an optimal cut-off (260 Du/L) determined by maximally selected rank statistics, and on the median value at T2 (250 Du/L). Dynamic changes between T0 and T1 were deemed meaningful if >10% of T1 or T0, whichever was greatest. Clinical outcome was estimated using the Kaplan-Meier method. Results: Median TKa (mTKa) at T0 was 73 Du/L (range 20-4302). As expected, P-containing treatment reduced mTKa levels at T1 (37 Du/L, range 20-4504). Conversely, at disease progression, TKa increased compared to T0 (mTKa at T2, 250 Du/L, range 20-3653). Median time to progression (mTTP) in pts with low TKa at T0 (n= 33) was 8.5 months, compared to 5.6 months in pts with high TKa (n= 12). Interestingly, pts with an increase in TKa at T1 (n=9) had a mTTP of only 3.1 months compared to pts with stable/reduced TKa (N=35), who showed a mTTP of 9 months. Considering the potential significance of TKa measured at disease progression, pts with high levels at T2 (n=18) had a worse outcome on subsequent post-study treatment (both chemotherapy and ET) compared to those with lower levels (n=18) (mTTP at T2, 2.9 vs 8.9 months, respectively). Conclusions: These data suggest for the first time that TKa may be a useful prognostic biomarker for non-invasive monitoring of MBC in the context of treatment with P. These results warrant further investigation in larger sample sets. Citation Format: Luca Malorni, Chiara Biagioni, Francesca De Luca, Martina Bonechi, Giuseppe Curigliano, Alessandro M. Minisini, Erica Moretti, Mattias Bergqvist, Karin Mattsson, Emanuela Risi, Ilenia Migliaccio, Stefania Vitale, Stefano Gabellini, Amelia McCartney, Irene De Santo, Francesca Galardi, Giulia Boccalini, Matteo Benelli, Lorenzo Rossi, Laura Biganzoli, Angelo Di Leo. Plasma thymidine kinase activity in patients with luminal metastatic breast cancer treated with Palbociclib within the phase II TREnd trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4416.

At the turn of the twentieth century, many Italian intellectuals opposed women’s participation in the public sphere, maintaining that women could not engage in politics due to their exclusive love for their biological children. Contemporary feminists countered this notion by promoting the idea of social motherhood. Sibilla Aleramo and Maria Montessori, better known for their work in feminist literature and early childhood education, respectively, made important contributions to this debate by implementing, theorizing, and popularizing the notion of social motherhood. This essay traces the strategies the two intellectuals used to demonstrate how women could act as political subjects via a socialization of the maternal functions. In her novel Una donna, Aleramo offered a fictional portrait of the social mother. Influenced by it and by the feminist debate on motherhood, Montessori conceptualized the notion of social motherhood as both a socialization of maternal duties and the expansion of women’s ma...

Aim: Anthracyclines are commonly used in breast cancer, although they lack validated predictive biomarkers. We explored the interaction between TOP2A protein by quantitative immunofluorescence (QIF) and anthracycline sensitivity. Patients & methods: Patients with estrogen receptor-negative breast cancer received neoadjuvant epirubicin. Pretreatment biopsies were analyzed using AQUA®. Total, cytoplasmic (C) and nuclear (N) TOP2A protein concentrations were expressed as QIF scores and compared with pathologic complete response (pCR), TOP2A by immunohistochemistry, TOP2A mRNA, TOP2A and HER2 gene status, and Ki-67 level. Results: In total, 76 cases were assessable. C, N, and total scores did not correlate with pCR, or other markers. The N:C ratio differed significantly by HER2 status. No pCRs occurred in patients in the lowest N:C quartile. Conclusion: Although no relevant correlation between TOP2A QIF scores and pCR was found, N:C ratio may have a negative predictive role, and may merit further exploration in a multifactorial predictive model that includes tumor and host factors.

Triple negative breast cancers, which are defined by lack of expression of estrogen, progesterone, or HER2 receptors, represent approximately 15% of all breast cancers, although they account for a much higher proportional of breast cancer mortality. This is due both to their innate aggressive biological characteristics, but also to lack of effective therapies. Conventional chemotherapy is currently the only treatment option, thus there is a critical need to find new and effective targeted therapies in this disease. While investigation of agents such as poly (ADP-ribose) polymerase (PARP) inhibitors and EGFR inhibitors continues, results from recent clinical trials indicate that these therapies are not as active in sporadic triple negative breast cancers as initially hoped. It is important therefore to consider other emerging therapeutic agents. Mutation in p53 is found in the vast majority of triple negative breast cancers, and as such is a target of particular interest. Within this review, several agents with potential activity against aberrant p53 signaling have been considered, as a novel approach to finding an effective targeted therapy for this aggressive breast cancer subtype.