Search

Background: The identification of the most appropriate targeted therapies for advanced cancers is challenging. We performed a molecular profiling of metastatic solid tumors utilizing a comprehensive next-generation sequencing (NGS) assay to determine genomic alterations' type, frequency, actionability, and potential correlations with PD-L1 expression., Methods: A total of 304 adult patients with heavily pretreated metastatic cancers treated between January 2019 and March 2021 were recruited. The CLIA-/UKAS-accredit Oncofocus assay targeting 505 genes was used on newly obtained or archived biopsies. Chi-square, Kruskal-Wallis, and Wilcoxon rank-sum tests were used where appropriate. Results were significant for P < .05., Results: A total of 237 tumors (78%) harbored potentially actionable genomic alterations. Tumors were positive for PD-L1 in 68.9% of cases. The median number of mutant genes/tumor was 2.0 (IQR: 1.0-3.0). Only 34.5% were actionable ESCAT Tier I-II with different prevalence according to cancer type. The DNA damage repair (14%), the PI3K/AKT/mTOR (14%), and the RAS/RAF/MAPK (12%) pathways were the most frequently altered. No association was found among PD-L1, ESCAT, age, sex, and tumor mutational status. Overall, 62 patients underwent targeted treatment, with 37.1% obtaining objective responses. The same molecular-driven treatment for different cancer types could be associated with opposite clinical outcomes., Conclusions: We highlight the clinical value of molecular profiling in metastatic solid tumors using comprehensive NGS-based panels to improve treatment algorithms in situations of uncertainty and facilitate clinical trial recruitment. However, interpreting genomic alterations in a tumor type-specific manner is critical., (© The Author(s) 2024. Published by Oxford University Press.)

Background: Neoadjuvant chemotherapy (NACT) is widely used in the treatment of triple-negative and HER2-positive breast cancer (BC), but its use in estrogen receptor (ER) and/or progesterone receptor (PR) positive/HER2-negative BC is questioned because of the low pathologic complete response (pCR) rates. This retrospective study assessed the mRNA-based MammaTyper® assay's capability of predicting pCR with NACT, and ER, PR, Ki67, and HER2 status at immunohistochemical (IHC) through transcriptomics., Methods: Diagnostic biopsies from 76 BC patients treated at the Cremona Hospital between 2012-2018 were analyzed. Relative mRNA expression levels of ERBB2, ESR1, PGR, and MKI67 were measured using the MammaTyper® kit and integrated into a pCR score. Predicting capability of pCR and standard IHC biomarkers could be assessed with ROC curves in 75 and 76 patients, respectively., Results: Overall, 68.0% patients obtained a MammaTyper® high-score and 32.0% a MammaTyper® low-score. Among high-score patients, 62.7% achieved pCR, compared to 16.7% in the low-score group (p = 0.0003). The binary MammaTyper® score showed good prediction of pCR in the overall cohort (area under curve [AUC] = 0.756) and in HR+/HER2-negative cases (AUC = 0.774). In cases with residual disease, the continuous MammaTyper® score correlated moderately with residual tumor size and decrease in tumor size. MammaTyper® showed substantial agreement with IHC for ESR1/ER and ERBB2/HER2, and moderate agreement for PGR/PR and MKI67/Ki67., Conclusion: Overall, MammaTyper® pCR score may serve as a standardized tool for predicting NACT response in HR+/HER2-negative BC, potentially guiding treatment strategies. Additionally, it could provide a more standardized and reproducible assessment of ER, PR, HER2, and Ki67 status., Competing Interests: Declaration of competing interest Francesco Schettini reports honoraria from Novartis, Gilead and Daiichy-Sankyo for educational events/materials and travel expenses from Novartis, Gilead and Daiichy-Sankyo. Daniele Generali declares personal fees for educational events by Novartis, Lilly, Pfizer, Daiichy-Sankyo, Roche; research funds from Astrazeneca, Novartis and LILT. Uğur Şahin is CEO and Mark Laible is an employee of BioNTech SE. Ralph M Wirtz is an employee of STRATIFYER Molecular Pathology GmbH. The other authors have nothing to declare., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Mathematical models based on partial differential equations (PDEs) can be exploited to handle clinical data with space/time dimensions, e.g. tumor growth challenged by neoadjuvant therapy. A model based on simplified assessment of tumor malignancy and pharmacodynamics efficiency was exercised to discover new metrics of patient prognosis in the OLTRE trial. We tested in a 17-patients cohort affected by early-stage triple negative breast cancer (TNBC) treated with 3 weeks of olaparib, the capability of a PDEs-based reactive-diffusive model of tumor growth to efficiently predict the response to olaparib in terms of SUV max detected at 18 FDG-PET/CT scan, by using specific terms to characterize tumor diffusion and proliferation. Computations were performed with COMSOL Multiphysics. Driving parameters governing the mathematical model were selected with Pearson's correlations. Discrepancies between actual and computed SUV max values were assessed with Student's t test and Wilcoxon rank sum test. The correlation between post-olaparib true and computed SUV max was assessed with Pearson's r and Spearman's rho. After defining the proper mathematical assumptions, the nominal drug efficiency (ε PD ) and tumor malignancy (r c ) were computationally evaluated. The former parameter reflected the activity of olaparib on the tumor, while the latter represented the growth rate of metabolic activity as detected by SUV max . ε PD was found to be directly dependent on basal tumor-infiltrating lymphocytes (TILs) and Ki67% and was detectable through proper linear regression functions according to TILs values, while r c was represented by the baseline Ki67-to-TILs ratio. Predicted post-olaparib SUV* max did not significantly differ from original post-olaparib SUV max in the overall, gBRCA-mutant and gBRCA-wild-type subpopulations (p > 0.05 in all cases), showing strong positive correlation (r = 0.9 and rho = 0.9, p < 0.0001 both). A model of simplified tumor dynamics was exercised to effectively produce an upfront prediction of efficacy of 3-week neoadjuvant olaparib in terms of SUV max . Prospective evaluation in independent cohorts and correlation of these outcomes with more recognized efficacy endpoints is now warranted for model confirmation and tailoring of escalated/de-escalated therapeutic strategies for early-TNBC patients., (© 2023. The Author(s).)

Introduction: Olaparib is effective in metastatic triple negative breast cancer (TNBC) carrying germline mutations in DNA damage repair (DDR) genes BRCA1/2 (g BRCA -mut). The OLTRE window-of-opportunity trial preliminarily investigated potential pathologic, radiometabolic and immune biomarkers of early-response to olaparib in g BRCA -wild-type (wt) TNBC and, as proof-of-concept in g BRCA -mut HER2-negative BC., Methods: Patients received olaparib for 3 weeks (3w) before standard neoadjuvant chemotherapy and underwent multiple FDG 18 -PET/CT scan (basal, after olaparib), clinical assessments (basal, every 3w), tumor biopsies and blood samplings (baseline, after olaparib). Clinical and radiometabolic responses were evaluated according to RECIST1.1 and PERCIST criteria., Results: 27 patients with g BRCA -wt TNBC and 8 with g BRCA -mut BC (6 TNBC, 2 HR+/HER2-negative) were enrolled. Three (11.1%) patients showed mutations in non- BRCA 1/2 DDR genes and 4 (14.8%) in other genes. 3w olaparib induced 16/35 and 15/27 partial clinical and radiometabolic responses, including in 40.7% and 50.0% g BRCA -wt patients. g BRCA -mut tumors presented numerically higher tumor-infiltrating lymphocytes (TILs) levels and PD-L1 positive tumors. Clinical responders experienced a reduction in T-regs/T-eff ratio (p=0.05), B and NK lymphocytes (p=0.003 both), with an average increase in T-helpers rate (p<0.001) and CD4/CD8 ratio (p=0.02). Ki67% and TILs did not vary significantly (p=0.67 and p=0.77). A numerical increase in PD-L1 positive cases after olaparib was observed, though non-significant (p=0.134). No differences were observed according to g BRCA status and type of response., Conclusions: Early-stage TNBC might be a target population for olaparib, irrespective of g BRCA mutations. Future trials should combine TILs, PD-L1 and g BRCA status to better identify candidates for escalated/de-escalated treatment strategies including olaparib., Competing Interests: EM, AB, LX, EZ and DC were employed by Personal Genomics Ltd. GW and ML are employed at Oncologica UK Ltd., which has received project funding from AstraZeneca outside of the submitted work. DG has declared consulting fees from Novartis, Lilly and Pfizer, research funding from LILT, Novartis, Astra-Zeneca and University of Trieste outside of the submitted work. IP has declared consulting fees from Roche, Novartis, Lilly, Pfizer, Astra-Zeneca, Pierre Fabre and Ipsen outside of the submitted work. GS has declared Grant/Research Support from MSD Italia S.r.l., consulting role for TESARO Bio Italy S.r.l. Johnson & Johnson and Clovis Oncology Italy S.r.l., outside of the submitted work. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Schettini, Corona, Giudici, Strina, Sirico, Bernocchi, Milani, Ziglioli, Aguggini, Azzini, Barbieri, Cervoni, Cappelletti, Molteni, Lazzari, Ferrero, Ungari, Marasco, Bruson, Xumerle, Zago, Cerra, Loddo, Williams, Paris, Scambia and Generali.)

Abstract: The novel coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is rapidly spreading throughout the world. The study describes 12 patients with SARS-CoV-2 pneumonia, who developed an acute erythematous rash with nonfollicular pinhead-sized pustules, without mucosal involvement. The clinical differential diagnosis was viral rash, acute generalized exanthematous pustulosis (AGEP), or multiform erythema. computed tomography with a diagnosis of interstitial pneumonia and a respiratory tract sample positive for SARS-CoV-2 in a reverse transcriptase polymerase chain reaction assay. Patients had signs of respiratory distress and were treated with hydroxychloroquine, darunavir, ritonavir, heparin, ceftriaxone, and azithromycin. Punch biopsies showed subcorneal pustules typical of AGEP. Dermal microvascular injury and thrombosis as described in skin damage by SARS-CoV-2 infection was not observed. The direct immunofluorescence for IgG, IgA, IgM, and C3 was negative in 8 patients investigated. A polymerase chain reaction for RNA SARS-CoV-2 performed on frozen skin was negative in 5 of 6 patients. Most of our patients were treated with systemic corticosteroids. After some days (4-10), the diffuse erythema and pustules had improved. AGEP is classified as a severe cutaneous adverse reaction, provoked by drugs and acute infections. Characteristically, removal of the offending agent leads to spontaneous resolution typically in less than 15 days. The recognition of AGEP is important, in order to avoid confusion with a systemic infection and consequently to avoid incorrect treatment. Cutaneous adverse reactions to drugs are common and are major health problems worldwide causing considerable costs for health care systems. We suggest that in the patients with AGEP during SARS-CoV-2 pneumonia, viral infection is a risk factor for developing drug reaction., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Introduction: Neoadjuvant hormonal therapy is generally considered a valid option for hormone receptor positive breast cancer (BC) patients who are unfit for chemotherapy or surgery., Aims: Whilst numerous studies analyzed efficacy of neoadjuvant chemotherapy (CT) or endocrine therapy (HT) alone in hormone receptor positive patients, there is a lack of research looking at the usefulness of a preoperative combinatorial approach of CT and HT in this patient subgroup., Methods: Using a predictive model previously described in the literature, developed to analyze the probability of benefit from preoperative chemotherapy, we were able to compare pathological complete response (pCR) rates expected with the use of CT alone with the pCR rates reported in a population of 192 patients treated with the combination of tamoxifen plus anthracycline-based CT at Cremona Hospital between 2003 and 2006., Results: Even with a relatively small patient population, this approach provided insightful information for the selection of hormone receptor positive BC patients most likely to benefit from the use of preoperative HT and CT in combination. Whilst no statistically significant benefit was obtained with the addition of tamoxifen to neoadjuvant chemotherapy in the entire population, or in any of the molecular stratification subgroups, the analysis of the calibration curve showed that a combinatorial approach may improve pCR in patients with luminal B tumors. More specific trials should be designed to confirm our initial results., Conclusion: To the best of our knowledge, this is the first report investigating the efficacy of the combination of CT and HT in the neoadjuvant treatment of hormone receptor positive BC.

Aim: Treatment of triple-negative breast cancer (TNBC) imposes great challenges, due to a lack of molecular targets. While use of gonadotropin-releasing hormone (GnRH) analogs has been validated in ER-positive breast cancer, this option has not been investigated in TNBC, even though a significant portion of these tumors upregulate GnRH receptors. We performed a meta-analysis of the literature to evaluate the effect of GnRH analogs in TNBC., Methods: Four studies were included in this study., Results: We detected a non-significant improvement in overall survival with GnRH analogs, while progression-free survival was unchanged., Discussion: The majority of the trials evaluated in this analysis were designed to test efficacy of GnRH analogs in preventing premature ovarian failure. This may represent a limitation of our study as these trials were not specifically designed to detect differences in survival outcome measures., Conclusion: Our results suggest that GnRH analogs may be useful as a targeted therapy in TNBC. Randomized prospective clinical trials are needed to investigate this hypothesis in the clinic.

Purpose: To identify hypoxia-related biomarkers indicative of response and resistance to epirubicin treatment in patients with locally advanced breast cancer., Patients and Methods: One hundred seventy-six women with T2-4 N0-1 breast tumours were randomly assigned to receive epirubicin 120 mg/m2/1-21 (EPI ARM), epirubicin 120 mg/m2/1-21 + erythropoietin 10.000 IU sc three times weekly (EPI-EPO ARM) and epirubicin 40 mg/m2/w-q21 (EPI-W ARM). Sixteen tumour proteins involved in cell survival, hypoxia, angiogenesis and growth factor, were assessed by immunohistochemistry in pre-treatment samples. A multivariate generalized linear regression approach was applied using a penalized least-square minimization to perform variable selection and regularization., Results: VEGF and GLUT-1 expression were significantly positively associated with complete response (CR) to treatment in all leave-one-out iterations. Bcl-2 expression was inversely correlated with pCR, whilst EPO expression was positively correlated with pathological complete response (pCR). Haemaglobin and HIF-1a nuclear expression were inversely correlated with pCR. HB and HIF-1a expression were associated with a higher risk of relapse and overall survival., Conclusion: Hypoxic biomarkers determines the epirubicin resistance in breast cancer. Assessment of such biomarkers, may be useful for predicting chemosensitivity and also anthracycline-based treatment outcome., Competing Interests: CONFLICTS OF INTEREST No declares that any conflicts of interest on this paper.

The OLTRE trial (ClinicalTrials.gov number: NCT02681562) is an open-label, 'window of opportunity' Phase II controlled trial to evaluate the biological activity of olaparib in locally advanced triple-negative breast cancer compared with other subtypes of locally advanced breast cancer patients carrying germinal BRCA mutation receiving olaparib with the same treatment approach. The primary end point is to investigate the correlation between baseline gene and protein expression profile in order to identify possible predictive markers of response to olaparib. The OLTRE trial is expected to identify the surrogate markers of the biological activity of olaparib in the treatment of patients with triple-negative breast cancer.

Introduction: Recently the combination of the mammalian target of rapamycin (mTOR) inhibitor everolimus and the aromatase inhibitor exemestane has been shown to double the progression-free survival rate in advanced breast cancer. However, the effect of the interrelated pathways of hypoxia-inducible factor-1α (HIF-1α) and mTOR signaling, both of which are associated with a more aggressive breast cancer phenotype and endocrine resistance, on response in the neoadjuvant setting is unknown. We, therefore, have investigated the influence of these pathways with the aim of better defining those patients most likely to benefit from an endocrine-based therapy associated with/without mTOR inhibitors., Patients and Methods: A total of 107 women with T2-4 N0-1 and estrogen receptor-positive breast cancer were randomly assigned to 6 months of primary letrozole (2.5 mg/daily) (LET) or LET plus oral "metronomic" cyclophosphamide (50mg/daily) (LET-CYC). Phospo-mTOR and HIF-1α were evaluated in tumor specimens collected before and after treatment using a tissue microarray format., Results: LET-based therapy induced a downregulation of phospho-mTOR and HIF-1α expression (P = .0001 and P < .004, respectively). The reduction of HIF-1α expression observed was positively correlated with phospho-mTOR reduction (P < .03); however, no treatment interaction between the two proteins was detected. HIF-1α expression was significantly modulated by the treatment (P < .004) with a reduction both in the LET arm (45%, n = 36/80) (P = .05) and LET-CYC arm (55%, n = 44/80) (P = .04). HIF-1α reduction showed a relationship with clinical response confined in LET arm only (P < .03)., Conclusions: In this neoadjuvant population, LET was able to modulate the phospho-mTOR and HIF-1α pathways and may define a subpopulation of nonresponders who may be most likely to benefit from mTOR inhibitors., (© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Purpose: To assess whether the combination of letrozole, metronomic cyclophosphamide and sorafenib (LCS) is well tolerated and shows activity in primary breast cancer (BC)., Methods: Thirteen oestrogen receptor-positive, postmenopausal, T2-4, N0-1 BC patients received the LCS combination for 6 months. In these patients we examined the pharmacokinetics of sorafenib and cyclophosphamide, toxicity of the regimen, the clinical response to therapy and changes in the levels of biologically relevant biomarkers., Results: Adequate plasma concentrations of sorafenib were achieved in patients when it was dosed in combination with L+C. The mean plasma concentrations of C were consistently lower following administration of LCS, compared with administration of L+C only. The most common drug-related grade 3/4 adverse events were skin rash (69.3%), hand-foot skin reaction (69.3%) and diarrhoea (46.1%). According to RECIST Criteria, a clinical complete response was observed in 6 of 13 patients. A significant reduction in tumour size, evaluated with MRI, was also observed between baseline and 14 days of treatment in all 13 patients (P=0.005). A significant reduction in SUV uptake, measured by (18)FDG-PET/CT, was observed in all patients between baseline and 30 days of treatment (P=0.015) and between baseline and definitive surgery (P=0.0002). Using modified CT Criteria, a response was demonstrated in 8 out of 10 evaluable patients at 30 days and in 11 out of 13 evaluable patients at the definitive surgery. A significant reduction in Ki67 expression was observed in all patients at day 14 compared with baseline (P<0.00001) and in 9 out of 13 patients at the definitive surgery compared with baseline (P<0.03). There was also a significant suppression of CD31 and VEGF-A expression in response to treatment (P=0.01 and P=0.007, respectively)., Conclusions: The LCS combination is feasible and tolerable. The tumour response and target biomarker modulation indicate that the combination is clinically and biologically active.

Introduction: The purpose of the present study was to investigate the relationship of expression of hypoxia inducible factor (HIF)-1α-modifying enzymes prolyl hydroxylase (PHD)1, PHD2 and PHD3 to response of tumours and survival in breast cancer patients enrolled in a phase II trial of neoadjuvant anthracycline and tamoxifen therapy., Methods: The expression of PHD1, PHD2 and PHD3 together with HIF-1α and the HIF-inducible genes vascular endothelial cell growth factor (VEGF) and carbonic anhydrase IX were assessed by immunohistochemistry using a tissue microarray approach in 211 patients with T2-4 N0-1 breast cancer enrolled in a randomised trial comparing single-agent epirubicin versus epirubicin and tamoxifen as the primary systemic treatment., Results: PHD1, PHD2 and PHD3 were detected in 47/179 (26.7%), 85/163 (52.2%) and 69/177 (39%) of tumours at baseline. PHD2 and PHD3 expression was moderate/strong whereas PHD1 expression was generally weak. There was a significant positive correlation between HIF-1α and PHD1 (P = 0.002) and PHD3 (P < 0.05) but not PHD2 (P = 0.41). There was a significant positive relationship between VEGF and PHD1 (P < 0.008) and PHD3 (P = 0.001) but not PHD2 (P = 0.09). PHD1, PHD2 and PHD3 expression was significantly increased after epirubicin therapy (all P < 0.000) with no significant difference in PHD changes between the treatment arms. There was no significant difference in response in tumours that expressed PHDs and PHD expression was not associated with survival., Conclusions: Although expression of the PHDs was not related to response or survival in patients receiving neoadjuvant epirubicin, our data provide the first evidence that these enzymes are upregulated on therapy in breast cancer and that the biological effects independent of HIF make them therapeutic targets.

Vascular endothelial growth factor A (VEGF-A) and vascular endothelial growth factor receptor 2 (VEGFR2) are the key factors mediating neo-vascularization. They are often coexpressed in breast cancer. Sex steroids may stimulate angiogenesis via the estrogen receptor (ER) pathway. We investigated to compare the effects of the addition of tamoxifen to epirubicin versus epirubicin alone on VEGF and VEGFR2 expression in breast cancer patients. The expression of VEGF and VEGFR2 was assessed on tissue microarray by immunohistochemistry at baseline conditions and after treatments in the case of 191 patients with T2-4 N0-1 breast cancer enrolled in a randomized trial comparing four cycles of single agent epirubicin versus epirubicin plus tamoxifen as primary systemic treatment. Epirubicin alone failed to induce changes in VEGF expression (P = 0.54), while the addition of tamoxifen to epirubicin resulted in a significant reduction in VEGF expression (P < 0.001). As a consequence, baseline VEGF had a negative prognostic role in patients who received epirubicin alone but not in patients receiving epirubicin plus tamoxifen (interaction test P < 0.05). VEGFR2 expression increased at residual tumor histology in both treatment arms, with a lesser extent in patients receiving tamoxifen plus epirubicin. Decrease in VEGFR2 expression was significantly associated with response rate (P = 0.02). The addition of tamoxifen to epirubicin resulted in a suppression of a key angiogenic pathway. These data suggest a potential synergism of these two drugs.

Purpose: We have shown previously that tumor infiltration by FOXP3+ regulatory T cells (Treg) is associated with increased relapse and shorter survival of patients with both in situ and invasive breast cancer. Because estrogen regulates Treg numbers in mice and promotes the proliferation of human Tregs, we hypothesized that blocking estrogen receptor-alpha signaling would abrogate Tregs and be associated with response to hormonal therapy and increased survival., Experimental Design: FOXP3+ Tregs were quantified in tumor samples collected at baseline by incisional biopsy and after 6 months at definitive surgery in 83 elderly breast cancer patients (T2-4 N0-1) enrolled in a randomized phase II trial based on 6 months of primary letrozole (2.5 mg/d) or 6 months of letrozole plus oral "metronomic" cyclophosphamide (50 mg/d)., Results: Treg number ranged from 0 to 380 (median, 30) before treatment and from 0 to 300 (median, 8) after treatment. There was a significant reduction in Tregs in letrozole and letrozole-cyclophosphamide patients (P < 0.0001 and P < 0.002, respectively) after treatment. Treg number at residual histology was inversely related with response (P < 0.03 and P = 0.50, respectively) and a greater Treg reduction was observed in responding patients (P < 0.03)., Conclusion: This study suggests that aromatase inhibitors may have an indirect antitumor mechanism of action through reducing Tregs in breast tumors and may be of use in estrogen receptor-alpha-negative tumors in combination with immunotherapy approaches.

Purpose: The phosphatidylinositol 3'-kinase (PI3K)/AKT/molecular target of rapamycin (mTOR) pathway is involved in the development of tumor resistance to endocrine therapy in breast cancer cell lines and represents an attractive target for pharmacologic intervention. However, the effects of endocrine therapy with aromatase inhibitors on in vivo expression of this signaling cascade, and its relation to tumor response and patient outcome, is unknown., Experimental Design: PI3K, phospho-AKT (pAKT) and phospho-mTOR were assessed by immunohistochemistry on tumor specimens collected at baseline and after 6 months of treatment in 113 elderly breast cancer patients consecutively enrolled in a randomized phase II trial of primary letrozole therapy and letrozole associated with metronomic cyclophosphamide., Results: Basal expression of the pathway was not significantly correlated with response or patient outcome. Both letrozole alone and letrozole with cyclophosphamide resulted in a significant reduction of PI3K expression (P = 0.02 and P < 0.005, respectively) and phospho-mTOR expression (P = 0.0001 and P = 0.0001, respectively). pAKT showed no change in the letrozole arm, whereas it was significantly decreased in the letrozole plus cyclophosphamide arm (P < 0.005). pAKT expression reduction was associated with a greater response rate (P = 0.05) and greater reduction in Ki67 expression (P = 0.05). Phospho-mTOR expression reduction was associated with a significantly longer disease-free survival in a multivariate analysis (P = 0.02)., Conclusions: Letrozole inhibits key molecules in the PI3K pathway that are important targets of new drugs being developed to overcome resistance. Changes in these molecules may have prognostic significance. These results should be taken into account when planning prospective trials testing up-front aromatase inhibitor with drugs targeting the PI3K/AKT/mTOR signaling pathway.