Early Modulation of Circulating MicroRNAs Levels in HER2-Positive Breast Cancer Patients Treated with Trastuzumab-Based Neoadjuvant Therapy.

Circulating microRNA (ct-miRNAs) are able to identify patients with differential response to HER2-targeted therapy. However, their dynamics are largely unknown. We assessed 752 miRNAs from 52 NeoALTTO patients with plasma pairs prior and two weeks after trastuzumab. Increased levels of ct-miR-148a-3p and ct-miR-374a-5p were significantly associated with pathological complete response (pCR) (p = 0.008 and 0.048, respectively). At a threshold ≥ the upper limit of the 95%CI of the mean difference, pCR resulted 45% (95%CI 24%-68%), and 44% (95%CI 22%-69%) for ct-miR-148a-3p and ct-miR-374a-5p, respectively. Notably, ct-miR-148a-3p retained its predictive value (OR 3.42, 95%CI 1.23-9.46, p = 0.018) in bivariate analysis along with estrogen receptor status. Combined information from ct-miR-148a-3p and ct-miR140-5p, which we previously reported to identify trastuzumab-responsive patients, resulted in greater predictive capability over each other, with pCR of 54% (95%CI 25%-81%) and 0% (95%CI 0%-31%) in ct-miR-148a/ct-miR-140-5p high/present and low/absent, respectively. GO and KEGG analyses showed common enriched terms between the targets of these ct-miRNAs, including cell metabolism regulation, AMPK and MAPK signaling, and HCC progression. In conclusion, early modulated ct-miR-148-3p may inform on the functional processes underlying treatment response, integrate the information from already available predictive biomarkers, and identify patients likely to respond to single agent trastuzumab-based neoadjuvant therapy.

Neoadjuvant letrozole plus taselisib versus letrozole plus placebo in postmenopausal women with oestrogen receptor-positive, HER2-negative, early-stage breast cancer (LORELEI): a multicentre, randomised, double-blind, placebo-controlled, phase 2 trial.

BACKGROUND: Endocrine therapy-based neoadjuvant treatment for luminal breast cancer allows efficient testing of new combinations before surgery. The activation of the phosphatidylinositol-3-kinase (PI3K) pathway is a known mechanism of resistance to endocrine therapy. Taselisib is an oral, selective PI3K inhibitor with enhanced activity against PIK3CA-mutant cancer cells. The LORELEI trial tested whether taselisib in combination with letrozole would result in an increased proportion of objective responses and pathological complete responses. METHODS: In this multicentre, randomised, double-blind, parallel-cohort, placebo-controlled phase 2, study, we enrolled postmenopausal women (aged ≥18 years) with histologically confirmed, oestrogen receptor (ER)-positive, HER2-negative, stage I-III, operable breast cancer, from 85 hospitals in 22 countries worldwide. To be eligible, patients had have an Eastern Cooperative Oncology Group (ECOG) performance status 0-1, adequate organ function, and had to have evaluable tumour tissue for PIK3CA genotyping. Patients were randomly assigned (1:1) by means of a permuted block algorithm (block size of four) via an interactive voice or web-based response system, to receive letrozole (2·5 mg/day orally, continuously) with either 4 mg of oral taselisib or placebo (on a 5 days-on, 2 days-off schedule) for 16 weeks, followed by surgery. Randomisation was stratified by tumour size and nodal status. Site staff, patients, and the sponsor were masked to treatment assignment. Coprimary endpoints were the proportion of patients who achieved an objective response by centrally assessed breast MRI and a locally assessed pathological complete response in the breast and axilla (ypT0/Tis, ypN0) at surgery in all randomly assigned patients and in patients with PIK3CA-mutant tumours. Analyses were done in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, number NCT02273973, and is closed to accrual. FINDINGS: Between Nov 12, 2014, and Aug 12, 2016, 334 participants were enrolled and randomly assigned to receive letrozole and placebo (n=168) or letrozole and taselisib (n=166). Median follow-up was 4·9 months (IQR 4·7-5·1). The study met one of its primary endpoints: the addition of taselisib to letrozole was associated with a higher proportion of patients achieving an objective response in all randomly assigned patients (66 [39%] of 168 patients in the placebo group vs 83 [50%] of 166 in the taselisib group; odds ratio [OR] 1·55, 95% CI 1·00-2·38; p=0·049) and in the PIK3CA-mutant subset (30 [38%] of 79 vs 41 [56%] of 73; OR 2·03, 95% CI 1·06-3·88; p=0·033). No significant differences were observed in pathological complete response between the two groups, either in the overall population (three [2%] of 166 in the taselisib group vs one [1%] of 168 in the placebo group; OR 3·07 [95% CI 0·32-29·85], p=0·37) or in the PIK3CA-mutant cohort (one patient [1%) vs none [0%]; OR not estimable, p=0·48). The most common grade 3-4 adverse events in the taselisib group were gastrointestinal (13 [8%] of 167 patients), infections (eight [5%]), and skin-subcutaneous tissue disorders (eight [5%]). In the placebo group, four (2%) of 167 patients had grade 3 or worse vascular disorders, two (1%) had gastrointestinal disorders, and two (1%) patients had grade 3 or worse infections and infestations. There was no grade 4 hyperglycaemia and grade 3 cases were asymptomatic. Serious adverse events were more common in the taselisib group (eight [5%] patients with infections and seven [4%] with gastrointestinal effects) than in the placebo group (one [1%] patient each with grade 3 postoperative wound and haematoma infection, grade 4 hypertensive encephalopathy, grade 3 acute cardiac failure, and grade 3 breast pain). One death occurred in the taselisib group, which was not considered to be treatment-related. INTERPRETATION: The increase in the proportion of patients who achieved an objective response from the addition of taselisib to endocrine therapy in a neoadjuvant setting is consistent with the clinical benefit observed in hormone receptor-positive, HER2-negative, metastatic breast cancer. FUNDING: Genentech and F Hoffmann-La Roche.

HER2-enriched subtype as a predictor of pathological complete response following trastuzumab and lapatinib without chemotherapy in early-stage HER2-positive breast cancer (PAMELA): an open-label, single-group, multicentre, phase 2 trial.

BACKGROUND: HER2-positive breast cancer consists of four intrinsic molecular subtypes-luminal A, luminal B, HER2-enriched, and basal-like-and a normal-like subtype, with the HER2-enriched subtype having the highest activation of the EGFR-HER2 pathway. We aimed to test the hypothesis that patients with the HER2-enriched subtype benefit the most from dual HER2 blockade. METHODS: PAMELA is an open-label, single-group, phase 2 trial done in 19 hospitals in Spain. We recruited female patients aged at least 18 years with previously untreated, centrally confirmed HER2-positive, stage I-IIIA invasive breast cancer regardless of hormone receptor status. Patients were given lapatinib (1000 mg per day orally) and trastuzumab (loading dose of 8 mg/kg, followed by 6 mg/kg every 3 weeks intravenously) for 18 weeks; hormone receptor-positive patients were additionally given letrozole (2·5 mg per day orally; if menopausal) or tamoxifen (20 mg per day orally; if premenopausal). Surgery was done 1-3 weeks after the last dose of study treatment. Intrinsic molecular subtypes of tumour biopsy samples taken at baseline (day 0) and day 14 were determined with the PAM50 predictor. The primary outcome was the ability of the HER2-enriched subtype to predict pathological complete response at the time of surgery. The primary outcome was assessed in the evaluable population (ie, all patients who had initial tumour biopsy samples available and who underwent definitive surgery) and safety was assessed in all patients who received at least one part of study treatment. This study is registered with ClinicalTrials.gov, number NCT01973660, and is completed. FINDINGS: Between Oct 28, 2013, and Nov 26, 2015, we recruited 151 patients, of whom 14 (9%) discontinued treatment and 137 (91%) completed treatment as planned. At baseline, most patients had the HER2-enriched subtype (101 [67%]), followed by luminal A (22 [15%]), luminal B (16 [11%]), basal-like (nine [6%]), and normal-like (three [2%]) subtypes. At the time of surgery, 46 (30%, 95% CI 23-39) of 151 patients had pathological complete response in the breast. 41 (41%, 31-51) of 101 patients with the HER2-enriched subtype and five (10%, 4-23) of 50 patients with non-HER2-enriched subtypes achieved pathological complete response at the time of surgery (odds ratio 6·2, 95% CI 2·3-16·8; p=0·0004). INTERPRETATION: The HER2-enriched subtype can identify patients with HER2-positive breast cancer who are likely to benefit from dual HER2 blockade therapies. FUNDING: GlaxoSmithKline, Susan Komen Foundation, CERCA Programme-Generalitat de Catalunya, Banco Bilbao Vizcaya Argentaria Foundation, Pas a Pas, and the Breast Cancer Research Foundation.

Plasma miRNA Levels for Predicting Therapeutic Response to Neoadjuvant Treatment in HER2-positive Breast Cancer: Results from the NeoALTTO Trial.

PURPOSE: To investigate the potential of circulating-miRNAs (ct-miRNA) as noninvasive biomarkers to predict the efficacy of single/dual HER2-targeted therapy in the NeoALTTO study. EXPERIMENTAL DESIGN: Patients with plasma samples at baseline (T0) and/or after 2 weeks (T1) of treatment were randomized into training (n = 183) and testing (n = 246) sets. RT-PCR-based high-throughput miRNA profiling was employed in the training set. After normalization, ct-miRNAs associated with pathologic complete response (pCR) were identified by univariate analysis. Multivariate logistic regression models were implemented to generate treatment-specific signatures at T0 and T1, which were evaluated by RT-PCR in the testing set. Event-free survival (EFS) according to ct-miRNA signatures was estimated by Kaplan-Meier method and Cox regression model. RESULTS: In the training set, starting from 51 ct-miRNAs associated with pCR, six signatures with statistically significant predictive capability in terms of area under the ROC curve (AUC) were identified. Four signatures were confirmed in the testing set: lapatinib at T0 and T1 [AUC 0.86; 95% confidence interval (CI), 0.73-0.98 and 0.71 (0.55-0.86)], respectively; trastuzumab at T1 (0.81; 0.70-0.92); lapatinib + trastuzumab at T1 (0.67; 0.51-0.83). These signatures were confirmed predictive after adjusting for known variables, including estrogen receptor status. ct-miRNA signatures failed to correlate with EFS. However, the levels of ct-miR-140-5p, included in the trastuzumab signature, were associated with EFS (HR 0.43; 95% CI, 0.22-0.84). CONCLUSIONS: ct-miRNAs discriminate patients with and without pCR after neoadjuvant lapatinib- and/or trastuzumab-based therapy. ct-miRNAs at week two could be valuable to identify patients responsive to trastuzumab, to avoid unnecessary combination with other anti-HER2 agents, and finally to assist deescalating treatment strategies.

Survival outcomes of the NeoALTTO study (BIG 1-06): updated results of a randomised multicenter phase III neoadjuvant clinical trial in patients with HER2-positive primary breast cancer.

BACKGROUND: Lapatinib (L) plus trastuzumab (T) with weekly paclitaxel significantly increased the pathologic complete response (pCR) rate compared with the anti-human epidermal growth factor receptor 2 (HER2) agent alone plus paclitaxel. The event-free survival (EFS) and overall survival (OS) by the treatment arms L + T vs. T and L vs. T and the relationship between pCR and EFS/OS both in the whole study population and according to hormone receptor-negative and hormone receptor-positive cohorts after a median follow-up of 6.7 years were assessed. PATIENTS AND METHODS: Four hundred fifty-five patients with HER2-positive early breast cancer randomly received L 1500 mg/day (n = 154), T (common dose, n = 149) or L 1000 mg/day plus T (n = 152) for 6 weeks, followed by the assigned anti-HER2 treatment combined with paclitaxel weekly × 12. After surgery, patients received 3 cycles of fluorouracil, epirubicin and cyclophosphamide. The primary end-point was pCR (ypT0/is; for current analysis, it is ypT0/is ypN0), and the secondary end-points were EFS and OS. RESULTS: Six-year EFS rates were 67%, 67% and 74% with L, T and L + T, respectively (L vs T: hazard ratio [HR], 0.98 [95% confidence interval {CI}, 0.64-1.51; P = .93]; L + T vs T: HR, 0.81 [95% CI, 0.52-1.26; P = .35]). Six-Year OS rates were 82%, 79% and 85% for L, T and L + T, respectively (L vs T: HR, 0.85 [95% CI, 0.49-1.46; P = .56]; L + T vs T: HR, 0.72 [95% CI, 0.41-1.27; P = .26]). In landmark analyses, patients with a pCR had a significantly higher 6-year EFS (77% and 65%) and OS (89% and 77%) compared with those without a pCR for both overall and the hormone receptor-negative cohort. CONCLUSION: Achieving a pCR is important in HER2-positive disease and translates into better long-term outcome with regard to EFS and OS.

Palbociclib and ribociclib in breast cancer: consensus workshop on the management of concomitant medication.

Drug-drug interactions are of significant concern in clinical practice in oncology, particularly in patients receiving Cyclin-dependent kinase (CDK) 4/6 inhibitors, which are typically exposed to long-term regimens. This article presents the highlights from the 'First Workshop on Pharmacology and Management of CDK4/6 Inhibitors: Consensus about Concomitant Medications'. The article is structured into two modules. The educational module includes background information regarding drug metabolism, corrected QT (QTc) interval abnormalities, management of psychotropic drugs and a comprehensive review of selected adverse effects of palbociclib and ribociclib. The collaborative module presents the conclusions of the five working groups, each of which comprised five experts from different fields. From these conclusions positive lists of drugs for treating common comorbid conditions that can be safely administered concomitantly with palbociclib and/or ribociclib were developed.

Radiation-induced changes in gene expression involve recruitment of existing messenger RNAs to and away from polysomes.

Although ionizing radiation has been shown to influence gene transcription, little is known about the effects of radiation on gene translational efficiency. To obtain a genome-wide perspective of the effects of radiation on gene translation, microarray analysis was done on polysome-bound RNA isolated from irradiated human brain tumor cells; to allow for a comparison with the effects of radiation on transcription, microarray analysis was also done using total RNA. The number of genes whose translational activity was modified by radiation was approximately 10-fold greater than those whose transcription was affected. The radiation-induced change in a gene's translational activity was shown to involve the recruitment of existing mRNAs to and away from polysomes. Moreover, the change in a gene's translational activity after irradiation correlated with changes in the level of its corresponding protein. These data suggest that radiation modifies gene expression primarily at the level of translation. In contrast to transcriptional changes, there was considerable overlap in the genes affected at the translational level among brain tumor cell lines and normal astrocytes. Thus, the radiation-induced translational control of a subset of mRNAs seems to be a fundamental component of cellular radioresponse.

Inhibition of Akt by the alkylphospholipid perifosine does not enhance the radiosensitivity of human glioma cells.

Akt has been implicated as a molecular determinant of cellular radiosensitivity. Because it is often constitutively activated or overexpressed in malignant gliomas, it has been suggested as a target for brain tumor radiosensitization. To evaluate the role of Akt in glioma radioresponse, we have determined the effects of perifosine, a clinically relevant alkylphospholipid that inhibits Akt activation, on the radiosensitivity of three human glioma cell lines (U87, U251, and LN229). Each of the glioma cell lines expressed clearly detectable levels of phosphorylated Akt indicative of constitutive Akt activity. Exposure to a perifosine concentration that reduced survival by approximately 50% significantly reduced the level of phosphorylated Akt as well as Akt activity. Cell survival analysis using a clonogenic assay, however, revealed that this Akt-inhibiting perifosine treatment did not enhance the radiosensitivity of the glioma cell lines. This evaluation was then extended to an in vivo model using U251 xenografts. Perifosine delivered to mice bearing U251 xenografts substantially reduced tumor phosphorylated Akt levels and inhibited tumor growth rate. However, the combination of perifosine and radiation resulted in a less than additive increase in tumor growth delay. Thus, in vitro and in vivo data indicate that the perifosine-mediated decrease in Akt activity does not enhance the radiosensitivity of three genetically disparate glioma cell lines. These results suggest that, although Akt may influence the radiosensitivity of other tumor types, it does not seem to be a target for glioma cell radiosensitization.

Neoadjuvant buparlisib plus trastuzumab and paclitaxel for women with HER2+ primary breast cancer: A randomised, double-blind, placebo-controlled phase II trial (NeoPHOEBE).

AIM: The Neoadjuvant PI3K inhibition in HER2 OverExpressing Breast cancEr (NeoPHOEBE) trial evaluated the efficacy and safety of buparlisib, a pan-phosphatidylinositol 3-kinase (PI3K) inhibitor, plus trastuzumab and paclitaxel as neoadjuvant treatment for human epidermal growth factor receptor-2 positive (HER2+) breast cancer. METHODS: NeoPHOEBE was a neoadjuvant, phase II, randomised, double-blind study. Women with HER2+ breast cancer were randomised within two independent cohorts by PIK3CA mutation status and, in each cohort stratified by oestrogen receptor (ER) status to receive buparlisib or placebo plus trastuzumab (first 6 weeks) followed by buparlisib or placebo with trastuzumab and paclitaxel. Primary end-point was pathological complete response (pCR) rate; key secondary end-point was objective response rate (ORR) at 6 weeks. Exploratory end-points were evaluation of Ki67 levels and change in tumour infiltrating lymphocytes (TILs) in intermediate biopsies at day 15. RESULTS: Recruitment was suspended mainly due to liver toxicity after enrolment of 50 of the planned 256 patients. In each arm (buparlisib n = 25; placebo n = 25) 21 patients (84%) had wild type PIK3CA and 4 patients (16%) had mutant PIK3CA. Overall, pCR rate was similar between buparlisib and placebo arms (32.0% versus 40%; one-sided P = 0.811). A trend towards higher ORR (68.8% versus 33.3%; P = 0.053) and a significant decrease in Ki67 (75% versus 26.7%; P = 0.021) was observed in buparlisib versus placebo arm in the ER+ subgroup (Pinteraction = 0.03). CONCLUSIONS: Addition of the pan-PI3K inhibitor buparlisib to taxane-trastuzumab-based therapy in HER2+ early breast cancer was not feasible. However, the higher ORR and Ki67 reduction in the ER+, HER2+ subgroup indicates a potential role for PI3K-targeted therapy in this setting and may warrant further investigation with better-tolerated second-generation PI3K inhibitors. TRIAL REGISTRATION IDENTIFIER: NCT01816594.

RNA Sequencing to Predict Response to Neoadjuvant Anti-HER2 Therapy: A Secondary Analysis of the NeoALTTO Randomized Clinical Trial.

IMPORTANCE: In neoadjuvant trials, treatment of human epidermal growth factor receptor 2 (HER2)-positive breast cancers with dual HER2 blockade resulted in increased pathologic complete response (pCR) rates compared with each targeted agent alone. Amplification and/or overexpression of HER2 currently remains the only biomarker for therapeutic decisions, but it is insufficient to explain the heterogeneous response to anti-HER2 agents. OBJECTIVE: To investigate the ability of clinically and biologically relevant genes and gene signatures (GSs) measured by RNA sequencing to predict the efficacy of anti-HER2 agents. DESIGN, SETTING, AND PARTICIPANTS: The neoadjuvant NeoALTTO trial randomized 455 women with HER2-positive early-stage breast cancer to trastuzumab, lapatinib, or the combination for 6 weeks followed by the addition of weekly paclitaxel for 12 weeks, followed by 3 cycles of fluorouracil, epirubicin, and cyclophosphamide after surgery. The present substudy, which was planned in the NeoALTTO main protocol, evaluated the association of pretreatment gene expression levels defined using RNA sequencing with pCR and event-free survival (EFS). MAIN OUTCOMES AND MEASURES: Gene expression-based biomarkers using RNA sequencing were examined for their association with response to anti-HER2 therapy and long-term outcome. RESULTS: Sequencing data were available for 254 (56%) of the NeoALTTO participants (mean [SD] age of substudy participants, 48.8 [11.2] years). The expression of ERBB2/HER2 was the most significant predictor of pCR, followed by HER2-enriched subtype, ESR1, treatment arm, ER immunohistochemical analysis scores, Genomic Grade Index, immune, proliferation, and AKT/mTOR GSs. Adjusting for clinicopathological variables and treatment arms, ERBB2/HER2, HER2-enriched subtype, ESR1, and Genomic Grade Index remained significant. Immune GSs were associated with higher pCR only in the combination arm (odds ratio, 2.1; 95% CI, 1.2-4.0; interaction test P = .01), while the stroma GSs were significantly associated with higher pCR in the single arms and with lower pCR in the combination arm (odds ratio, 0.46; 95% CI, 0.25-0.84; P = .009). None of the evaluated variables was associated with EFS after correction for multiple testing, but this analysis was underpowered. CONCLUSIONS AND RELEVANCE: High levels of ERBB2/HER2 and low levels of ESR1 were associated with pCR in all treatment arms. In the combination arm, high expression of immune and stroma GSs were significantly associated with higher and lower pCR rates, respectively, and should be further explored as candidate predictive markers. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00553358.

PIK3CA mutations are associated with decreased benefit to neoadjuvant human epidermal growth factor receptor 2-targeted therapies in breast cancer.

PURPOSE: We investigated whether mutations in the gene encoding the phosphatidylinositol 3-kinase (PI3K) catalytic subunit (PIK3CA) correlates with response to neoadjuvant human epidermal growth factor receptor 2 (HER2) -targeted therapies in patients with breast cancer. PATIENTS AND METHODS: Baseline tissue biopsies were available from patients with HER2-positive early breast cancer who were enrolled onto the Neoadjuvant Lapatinib and/or Trastuzumab Treatment Optimization trial (NeoALTTO). Activating mutations in PIK3CA were identified using mass spectrometry-based genotyping. RESULTS: PIK3CA mutations were identified in 23% of HER2-positive breast tumors, and these mutations were associated with poorer outcome in all of the treatment arms. Patients treated with a combination of trastuzumab and lapatinib who had wild-type PIK3CA obtained a total pathologic complete response (pCR) rate of 53.1%, which decreased to 28.6% in patients with tumors that carried PIK3CA activating mutations (P = .012). CONCLUSION: Activating mutations in PIK3CA predicted poor pCR in patients with HER2-positive breast cancer treated with neoadjuvant therapies that target HER2. Consequently, the combination of anti-HER2 agents and PI3K inhibitors is being investigated.