Red Blood Cells Protein Profile Is Modified in Breast Cancer Patients.

Metastasis is the primary cause of death for most breast cancer (BC) patients who succumb to the disease. During the hematogenous dissemination, circulating tumor cells interact with different blood components. Thus, there are microenvironmental and systemic processes contributing to cancer regulation. We have recently published that red blood cells (RBCs) that accompany circulating tumor cells have prognostic value in metastatic BC patients. RBC alterations are related to several diseases. Although the principal known role is gas transport, it has been recently assigned additional functions as regulatory cells on circulation. Hence, to explore their potential contribution to tumor progression, we characterized the proteomic composition of RBCs from 53 BC patients from stages I to III and IV, compared with 33 cancer-free controls. In this work, we observed that RBCs from BC patients showed a different proteomic profile compared to cancer-free controls and between different tumor stages. The differential proteins were mainly related to extracellular components, proteasome, and metabolism. Embryonic hemoglobins, not expected in adults' RBCs, were detected in BC patients. Besides, lysosome-associated membrane glycoprotein 2 emerge as a new RBCs marker with diagnostic and prognostic potential for metastatic BC patients. Seemingly, RBCs are acquiring modifications in their proteomic composition that probably represents the systemic cancer disease, conditioned by the tumor microenvironment.

Noninvasive detection of microsatellite instability in patients with endometrial cancer.

The analysis of mismatch repair proteins in solid tissue is the standard of care (SoC) for the microsatellite instability (MSI) characterization in endometrial cancer (EC). Uterine aspirates (UAs) or circulating-DNA (cfDNA) samples capture the intratumor heterogeneity and provide a more comprehensive and dynamic molecular diagnosis. Thus, MSI analysis by droplet-digital PCR (ddPCR) in UAs and cfDNA can provide a reliable tool to characterize and follow-up the disease. The UAs, paraffin-embedded tumor tissue (FFPE) and longitudinal plasma samples from a cohort of 90 EC patients were analyzed using ddPCR panel and compared to the SoC. A high concordance (96.67%) was obtained between the analysis of MSI markers in UAs and the SoC. Three discordant cases were validated as unstable by ddPCR on FFPE samples. Besides, a good overall concordance (70.27%) was obtained when comparing the performance of the ddPCR assay on UAs and cfDNA in high-risk tumors. Importantly, our results also evidenced the value of MSI analysis to monitor the disease evolution. MSI evaluation in minimally invasive samples shows great accuracy and sensitivity and provides a valuable tool for the molecular characterization and follow-up of endometrial tumors, opening new opportunities for personalized management of EC.

Improving the Management of Endometrial Cancer Patients through the Use of Liquid Biopsy Analyses: A Case Report.

Endometrial cancer (EC) is the 4th most common neoplasm of the female genital tract, with 15-20% of patients being of high risk of recurrence which leads to a significant decrease in patient survival. Current therapeutic options for patients with EC are poor, being the combined therapy of carboplatin and paclitaxel the standard of care, with limited efficacy. Therefore, new therapeutic options and better monitoring tools are needed to improve the management of the disease. In the current case report, we showcase the value of liquid biopsy analyses in a microsatellite instability EC patient with initially good prognosis that however underwent rapid progression disease within 6 months post-surgery; through the study of plasma cfDNA/ctDNA dynamics to assess the tumour evolution during treatment, as well as the study of the uterine aspirate as a valuable sample that captures the intra-tumour heterogeneity that allows a comprehensive genomic profiling of the disease to identify potential therapeutic options. Furthermore, preclinical models were generated at the time of tumour progression to assess the efficacy of the identified targeted therapies.

Use of image analysis to evaluate morphometric measurements of broiler breast fillets affected by the woody breast condition.

The objective of this study was to investigate the relationships between morphometric measurements and woody breast (WB) severity in breast fillets using image processing as an objective detection method for WB. Breast fillets were collected and categorized as normal (NORM), mild (MILD), moderate (MOD), and severe (SEV). Compression force and energy increased as WB severity increased alongside a decrease in severity through fillet regions (P < 0.05). Length and caudal thickness were highest in SEV and MOD groups (P < 0.05), and cranial thickness increased as WB severity increased (P < 0.05). The aerial area was the smallest in NORM fillets, while the planar area increased from NORM to MOD (P < 0.05). Fillet curvatures were highest in SEV and MOD fillets (P < 0.05). All measured parameters expressed strong correlation to WB scores (P < 0.05) except width. The results from this study may provide a basis for further assessment of the potential incorporation of these measurements into vision grading systems that may allow processors to sort fillets by WB severity in commercial plants. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-022-05525-x.

Assessment of meat quality distributions of breast fillets with woody breast condition in the raw and cooked state.

The objective of this study was to determine meat quality distributions and assess hardness distributions in raw and cooked breast fillets with the woody breast (WB) condition, in addition to evaluating the relationship between water properties and WB severity. A total of 90 breast fillets were collected and categorized as normal (NORM), mild (MILD) and severe (SEV). Breast weight, drip loss, compression measurements, cook loss, shear and texture profile analysis (TPA) values were measured for each sample by fillet location (cranial to caudal) and sampling depth (cranial-superficial, cranial-internal, middle-superficial, and middle-internal) in the raw and cooked meat state. Low-field NMR relaxation measurements were also collected for both the raw and cooked fillets. Results indicate that severe WB expressed increased hardness, a higher water content (bound water and free water) and reduced meat quality attributes in raw and cooked meat. Breast fillet hardness and meat quality distributions were unevenly distributed between fillets, compression measurements were higher mainly in the cranial region, and progressively decreased toward the caudal region for both raw and cooked fillets. Shear force and energy values were higher in the cranial region than in the middle region, and TPA values were higher in superficial regions rather than internal portions. Additionally, low-field NMR could be used to predict WB through variation in water properties as thermal processing reduces water distributing abilities in affected fillets.

Quality properties of chicken meatballs prepared with varying proportions of woody breast meat.

The objective of this study was to explore the effect of woody breast (WB) on quality characteristics of chicken meatballs paired with the feasibility of its inclusion. Cook loss (CL), color (CIE L*, a*, b*), texture (hardness, springiness, chewiness and resilience), low-field NMR (bound water, immobilized water, and free water), microstructure, and sensory characteristics of chicken meatballs with different WB inclusion levels (0%, 25%, 50%, 75%, 100%) were analyzed. The results showed that the impairment of product quality traits such as CL, color, texture (hardness, chewiness), free water, microstructure, and sensory scores (appearance, organization, total score) increased as the percentage of WB meat increased in the product formulation, particularly when the WB incorporation level exceeded 25%. Indeed, cook loss, L*, a*, b* parameters, bound water, and immobilized water increased when the WB inclusion level was higher than 25% (P ≤ 0.05). However, free water, sensory characteristics, hardness, and chewiness parameters decreased (P ≤ 0.05). The microstructure of chicken meatballs also changed as the proportion of WB meat increased. Even though data suggest that the inclusion of WB meat up to 30% could be feasible to produce acceptable chicken meatballs, the optimal maximum incorporation rate of WB meat into chicken meatball recipes was 25% based on economic feasibility and final overall quality.

An olaparib window-of-opportunity trial in patients with early-stage endometrial carcinoma: POLEN study.

OBJECTIVE: Olaparib is a potent inhibitor of poly(ADP-ribose) polymerase (PARP)-1, 2, and 3 with potential activity in endometrial cancer (EC). METHODS: In this window-of-opportunity trial, women with operable type 1 EC received olaparib oral tablets (300mg) twice daily for 28days before surgery. The primary objective was to evaluate the effects of olaparib on EC in tissue samples taken at baseline and at treatment completion. Signal of activity was defined as significant changes in the expression of the cell cycle-related proteins cyclin D1, Ki67, and cleaved caspase-3. RESULTS: A total of 31 patients were included in the biomarker analysis. The median time of olaparib exposure was 24 days (1-39). Significant inhibition was found for cyclin D1 (p < 0.01), but not for Ki67 and active caspase 3 immunostaining. PARP-1 levels positively correlated with cyclin D1 levels (rho = 0.661, p = 0.0001). Both PARP-1 and cyclin D1 levels were significantly lower (p = 0.022 and p = 0.004, respectively) in patients with ARID1A[-] tumors than ARID1A[+] tumors. A significant relationship between plasma olaparib concentrations and decreased GLUT1 activity was observed (r = -0.5885; p < 0.05). Drug-related toxicity consisted mostly of gastrointestinal and grade 1 or 2 adverse events. CONCLUSIONS: Olaparib reduced expression of cyclin D1, which positively correlated with PARP-1 levels. This effect was more evident in ARID1A-deficient tumors. Olaparib further induced inhibition of GLUT1 plasma activity. Our findings could have noteworthy implications in predicting which patients with EC would benefit from olaparib-based strategies.

CTCs-derived xenograft development in a triple negative breast cancer case.

Triple-negative breast cancer (TNBC) is characterized by high rates of metastasis and no available molecular targets. CTCs derived xenografts (CDX) have demonstrated to be a promising tool for understanding cancer biology. In our study, a CDX from a TNBC patient was developed for the first time. After CDX characterization, WNT signaling was found as the main mechanism related with this tumor biology and potential CTCs markers were identified and subsequently validated in TNBC patients. In this cohort high levels of MELK expression were associated with poorer survival rates. Overall, our study demonstrates that CTCs from TNBC are tumorigenic and CDXs are a useful model to obtain valuable information about the tumor.

Identification of 526 conserved metazoan genetic innovations exposes a new role for cofactor E-like in neuronal microtubule homeostasis.

The evolution of metazoans from their choanoflagellate-like unicellular ancestor coincided with the acquisition of novel biological functions to support a multicellular lifestyle, and eventually, the unique cellular and physiological demands of differentiated cell types such as those forming the nervous, muscle and immune systems. In an effort to understand the molecular underpinnings of such metazoan innovations, we carried out a comparative genomics analysis for genes found exclusively in, and widely conserved across, metazoans. Using this approach, we identified a set of 526 core metazoan-specific genes (the 'metazoanome'), approximately 10% of which are largely uncharacterized, 16% of which are associated with known human disease, and 66% of which are conserved in Trichoplax adhaerens, a basal metazoan lacking neurons and other specialized cell types. Global analyses of previously-characterized core metazoan genes suggest a prevalent property, namely that they act as partially redundant modifiers of ancient eukaryotic pathways. Our data also highlights the importance of exaptation of pre-existing genetic tools during metazoan evolution. Expression studies in C. elegans revealed that many metazoan-specific genes, including tubulin folding cofactor E-like (TBCEL/coel-1), are expressed in neurons. We used C. elegans COEL-1 as a representative to experimentally validate the metazoan-specific character of our dataset. We show that coel-1 disruption results in developmental hypersensitivity to the microtubule drug paclitaxel/taxol, and that overexpression of coel-1 has broad effects during embryonic development and perturbs specialized microtubules in the touch receptor neurons (TRNs). In addition, coel-1 influences the migration, neurite outgrowth and mechanosensory function of the TRNs, and functionally interacts with components of the tubulin acetylation/deacetylation pathway. Together, our findings unveil a conserved molecular toolbox fundamental to metazoan biology that contains a number of neuronally expressed and disease-related genes, and reveal a key role for TBCEL/coel-1 in regulating microtubule function during metazoan development and neuronal differentiation.

Annexin-A2 as predictor biomarker of recurrent disease in endometrial cancer.

Endometrial carcinomas, the most common malignant tumour of the female genital tract, are usually diagnosed at an early stage with uterine-confined disease and an overall favourable prognosis. However, up to 20% of endometrial carcinomas will end up in recurrent disease, associated with a drop in survival and representing the major clinical challenge. Management of this group of risk patients relies on robust biomarkers that may predict which endometrial carcinomas will relapse. For this, we performed a proteomic analysis comparing primary lesions with recurrences and identified ANXA2 as a potential biomarker associated with recurrent disease that we further validated in an independent series of samples by immunohistochemistry. We demonstrated in vitro a role for ANXA2 in the promotion of metastasis rather than interfering with sensitivity to radio/chemotherapy. In addition, ANXA2 silencing resulted in a reduced metastatic pattern in a mice model of endometrial cancer dissemination, with a limited presence of circulating tumor cells. Finally, a retrospective study in a cohort of 93 patients showed that ANXA2 effectively predicted those endometrioid endometrial carcinomas that finally recurred. Importantly, ANXA2 demonstrated a predictive value also among low risk Stage I endometrioid endometrial carcinomas, highlighting the clinical utility of ANXA2 biomarker as predictor of recurrent disease in endometrial cancer. Retrospective and prospective studies are ongoing to validate ANXA2 as a potential tool for optimal stratification of patients susceptible to receive radical surgery and radio/chemotherapy.

Molecular profiling of circulating tumor cells links plasticity to the metastatic process in endometrial cancer.

BACKGROUND: About 20% of patients diagnosed with endometrial cancer (EC) are considered high-risk with unfavorable prognosis. In the framework of the European Network for Individualized Treatment in EC (ENITEC), we investigated the presence and phenotypic features of Circulating Tumor Cells (CTC) in high-risk EC patients. METHODS: CTC isolation was carried out in peripheral blood samples from 34 patients, ranging from Grade 3 Stage IB to Stage IV carcinomas and recurrences, and 27 healthy controls using two methodologies. Samples were subjected to EpCAM-based immunoisolation using the CELLection™ Epithelial Enrich kit (Invitrogen, Dynal) followed by RTqPCR analysis. The phenotypic determinants of endometrial CTC in terms of pathogenesis, hormone receptor pathways, stem cell markers and epithelial to mesenchymal transition (EMT) drivers were asked. Kruskal-Wallis analysis followed by Dunn's post-test was used for comparisons between groups. Statistical significance was set at p < 0.05. RESULTS: EpCAM-based immunoisolation positively detected CTC in high-risk endometrial cancer patients. CTC characterization indicated a remarkable plasticity phenotype defined by the expression of the EMT markers ETV5, NOTCH1, SNAI1, TGFB1, ZEB1 and ZEB2. In addition, the expression of ALDH and CD44 pointed to an association with stemness, while the expression of CTNNB1, STS, GDF15, RELA, RUNX1, BRAF and PIK3CA suggested potential therapeutic targets. We further recapitulated the EMT phenotype found in endometrial CTC through the up-regulation of ETV5 in an EC cell line, and validated in an animal model of systemic dissemination the propensity of these CTC in the accomplishment of metastasis. CONCLUSIONS: Our results associate the presence of CTC with high-risk EC. Gene-expression profiling characterized a CTC-plasticity phenotype with stemness and EMT features. We finally recapitulated this CTC-phenotype by over-expressing ETV5 in the EC cell line Hec1A and demonstrated an advantage in the promotion of metastasis in an in vivo mouse model of CTC dissemination and homing.

The doublecortin-related gene zyg-8 is a microtubule organizer in Caenorhabditis elegans neurons.

Doublecortin-domain containing (DCDC) genes play key roles in the normal and pathological development of the human brain cortex. The origin of the cellular specialisation and the functional redundancy of these microtubule (MT)-associated proteins (MAPs), especially those of Doublecortin (DCX) and Doublecortin-like kinase (DCLKs) genes, is still unclear. The DCX domain has the ability to control MT architecture and bundling. However, the physiological significance of such properties is not fully understood. To address these issues, we sought post-mitotic roles for zyg-8, the sole representative of the DCX-DCLK subfamily of genes in C. elegans. Previously, zyg-8 has been shown to control anaphase-spindle positioning in one-cell stage embryos, but functions of the gene later in development have not been investigated. Here we show that wild-type zyg-8 is required beyond early embryonic divisions for proper development, spontaneous locomotion and touch sensitivity of adult worms. Consistently, we find zyg-8 expression in the six touch receptor neurons (TRNs), as well as in a subset of other neuronal and non-neuronal cells. In TRNs and motoneurons, zyg-8 controls cell body shape/polarity and process outgrowth and morphology. Ultrastructural analysis of mutant animals reveals that zyg-8 promotes structural integrity, length and number of individual MTs, as well as their bundled organisation in TRNs, with no impact on MT architecture.

Zoledronic acid in the treatment of metastatic breast cancer.

Bone is the most common location of metastatic disease. Approximately 80% of all bone metastases are observed in patients with breast or prostate tumours and are responsible for more than 300 000 deaths every year. Treatment of malignant bone disease with bisphosphonates has been shown to reduce bone events and delay their onset, and several reviews and meta-analyses have confirmed the benefit of these drugs in controlling bone metastases. Zoledronic acid is a bisphosphonate that has been shown to delay or prevent the development of skeletal-related events in patients with bone metastases. Furthermore, compared with other bisphosphonates, zoledronic acid has also shown better pain control and various studies also suggest an improvement in quality of life, although with no impact on overall survival. The duration and optimal regimen for long-term zoledronic acid therapy have not yet been defined, but some studies suggest that continuing zoledronic acid therapy for more than 2 years could also extend its beneficial effect.

Preventing alpelisib-related hyperglycaemia in HR+/HER2-/PIK3CA-mutated advanced breast cancer using metformin (METALLICA): a multicentre, open-label, single-arm, phase 2 trial.

Background: Hyperglycaemia is an early and frequent adverse event during alpelisib treatment. METALLICA aimed to evaluate prophylactic metformin to prevent or reduce hyperglycaemia occurrence in patients with HR+/HER2-/PIK3CA-mutated advanced breast cancer (ABC). Methods: Between August 13th, 2020 and March 23rd, 2022, this 2-cohort, phase 2, multicentre, single-arm trial (NCT04300790) enrolled patients with HR+/HER2-/PIK3CA-mutated ABC: cohort A, normal glycaemia (fasting plasma glucose <100 mg/dL [<5.6 mmol/L] and HbA1c <5.7%), and cohort B, prediabetes (fasting plasma glucose 100-140 mg/dL [5.6-7.8 mmol/L] and/or haemoglobin A1C [HbA1c] 5.7-6.4%). Participants were at least 18 years old, with Eastern Cooperative Oncology Group performance status of 0-1, and up to two prior lines of endocrine therapy (ET) for ABC. Alpelisib plus ET were administered in 28-day cycles after initiation of prophylactic metformin plus ET. Primary endpoint was the incidence of grade 3-4 hyperglycaemia over the first 8 weeks. Secondary endpoints included safety, progression-free survival (PFS), objective response rate (ORR), and clinical benefit rate (CBR). The primary objective for cohort A and B is met with ≤7 (14.6%) and ≤4 (20%) patients with grade 3-4 hyperglycaemia over the first 8 weeks, respectively. Findings: 233 patients were screened, and 68 (20.2%) patients were enrolled in cohorts A (n = 48) and B (n = 20). Median follow-up was 7.8 months (IQR 1.4-19.6). Over the first 8 weeks, one (2.1%) of 48 patients in cohort A (95% CI: 0.5-11.1; P < 0.0001), and three (15.0%) of 20 patients in cohort B (95% CI: 5.6-37.8; P = 0.016) had grade 3-4 hyperglycaemia. Serious treatment-related adverse events occurred in seven patients (10.3%). The most common were rash (two [2.9%]), vomiting (two [2.9%]), and diarrhoea (two [2.9%]). Discontinuation of alpelisib caused by AEs was reported in nine patients (13.2%), none caused by hyperglycaemia. At data cutoff (15 June, 2022), no treatment-related deaths were observed. In the full analysis set, median PFS was 7.3 months (95% CI: 5.9-not reached), ORR was 20.6% (95% CI: 11.7-32.1%), and CBR was 52.9% (95% CI: 40.4-65.2). Interpretation: In HR+/HER2-/PIK3CA-mutated ABC, prophylactic metformin before alpelisib plus endocrine treatment has low incidence and severity of alpelicib-induced hyperglycaemia. Funding: Novartis Pharmaceuticals.

The major alpha-tubulin K40 acetyltransferase alphaTAT1 promotes rapid ciliogenesis and efficient mechanosensation.

Long-lived microtubules found in ciliary axonemes, neuronal processes, and migrating cells are marked by α-tubulin acetylation on lysine 40, a modification that takes place inside the microtubule lumen. The physiological importance of microtubule acetylation remains elusive. Here, we identify a BBSome-associated protein that we name αTAT1, with a highly specific α-tubulin K40 acetyltransferase activity and a catalytic preference for microtubules over free tubulin. In mammalian cells, the catalytic activity of αTAT1 is necessary and sufficient for α-tubulin K40 acetylation. Remarkably, αTAT1 is universally and exclusively conserved in ciliated organisms, and is required for the acetylation of axonemal microtubules and for the normal kinetics of primary cilium assembly. In Caenorhabditis elegans, microtubule acetylation is most prominent in touch receptor neurons (TRNs) and MEC-17, a homolog of αTAT1, and its paralog αTAT-2 are required for α-tubulin acetylation and for two distinct types of touch sensation. Furthermore, in animals lacking MEC-17, αTAT-2, and the sole C. elegans K40α-tubulin MEC-12, touch sensation can be restored by expression of an acetyl-mimic MEC-12[K40Q]. We conclude that αTAT1 is the major and possibly the sole α-tubulin K40 acetyltransferase in mammals and nematodes, and that tubulin acetylation plays a conserved role in several microtubule-based processes.

Real-world safety and effectiveness of maintenance niraparib for platinum-sensitive recurrent ovarian cancer: A GEICO retrospective observational study within the Spanish expanded-access programme.

AIM: To describe patient characteristics, effectiveness and safety in a real-world population treated with niraparib in the Spanish expanded-access programme. PATIENTS AND METHODS: This retrospective observational study included women with platinum-sensitive recurrent high-grade serous ovarian cancer who received maintenance niraparib within the Spanish niraparib expanded-access programme. Eligible patients had received ≥2 previous lines of platinum-containing therapy, remained platinum-sensitive after the penultimate line of platinum and had responded to the most recent platinum-containing therapy. Niraparib dosing was at the treating physician's discretion (300 mg/day fixed starting dose or individualised starting dose [ISD] according to baseline body weight and platelet count). Safety, impact of dose adjustments, patient characteristics and effectiveness were analysed using data extracted from medical records. RESULTS: Among 316 eligible patients, 80% had BRCA wild-type tumours and 66% received an ISD. Median niraparib duration was 7.8 months. The most common adverse events typically occurred within 3 months of starting niraparib. Median progression-free survival was 8.6 (95% confidence interval [CI] 7.6-10.0) months. One- and 2-year overall survival rates were 86% (95% CI 81-89%) and 65% (95% CI 59-70%), respectively. Dose interruptions, dose reductions, haematological toxicities and asthenia/fatigue were less common with ISD than fixed starting dose niraparib, but progression-free survival was similar irrespective of dosing strategy. Subsequent therapy included platinum in 71% of patients who received further treatment. CONCLUSION: Outcomes in this large real-world dataset of niraparib-treated patients are consistent with phase III trials, providing reassuring evidence of the tolerability and activity of niraparib maintenance therapy for platinum-sensitive recurrent ovarian cancer. GOV REGISTRATION: NCT04546373.

Open-label phase II clinical trial of ketoconazole as CYP17 inhibitor in metastatic or advanced non-resectable granulosa cell ovarian tumors: the GREKO (GRanulosa Et KetOconazole) trial, GETHI 2011-03.

BACKGROUND: Granulosa cell ovarian tumor (GCT) is characterized by a pathognomonic mutation in the FOXL2 gene (402 C > G) that leads to an overactivation of steroidogenesis. CYP17 is a key enzyme in such process and can be inhibited by ketoconazole. METHODS: We designed a phase II clinical trial to assess the efficacy of ketoconazole in advanced GCT and conducted several in vitro studies to support the clinical findings. RESULTS: From October 1st 2012 to January 31st 2014, six evaluable patients were recruited in ten hospitals of the Spanish Group for Transversal Oncology and Research in Orphan and Infrequent Tumors" (GETTHI). FOXL2 (402C > G) mutation was confirmed in three; two cases were wild type and it could not be assessed in one. No objective response by RECIST was observed, but five cases achieved stable disease longer than 12 months. Median progression-free survival was 14.06 months (CI 95% 5.43-22.69) for the whole study population (3.38 and 13.47 months for wild-type cases and 14.06, 20.67 and 26.51 for those with confirmed FOXL2 mutation). Median overall survival was 22·99 months (CI 95% 8.99-36.99). In vitro assays confirmed the activity of ketoconazole in this tumor and suggested potential synergisms with other hormone therapies. CONCLUSION: Ketoconazole has shown activity in advanced GCT in clinical and in vitro studies. Based on these data, an orphan designation was granted by the European Medicines Agency for ketoconazole in GCT (EU/3/17/1857). GOV IDENTIFIER: NCT01584297.

Methylphenidate in the management of asthenia in breast cancer patients treated with docetaxel: results of a pilot study.

BACKGROUND: The objectives of this pilot study were to evaluate the safety and efficacy of the central nervous system stimulant methylphenidate in the management of asthenia in breast cancer patients treated with docetaxel. PATIENTS AND METHODS: Patients with early breast cancer who presented asthenia >3 on the Visual Analogue Scale (VAS) after the first cycle of docetaxel-based chemotherapy were included. Patients received two additional cycles of chemotherapy, one with methylphenidate (10 mg bid) and the other without methylphenidate. Asthenia was evaluated using VAS and the Functional Assessment of Cancer Therapy-Fatigue (FACT-F) scale. Distress was assessed using the Hospital Anxiety and Depression Scale (HADS), and quality of life using FACT-F. RESULTS: Ten patients were included and evaluated for efficacy and safety. Overall, cycles with methylphenidate were better tolerated than those without methylphenidate in terms of asthenia (VAS, p = 0.004; FACT-F, p = 0.027) and quality of life (FACT-F, p = 0.047). No significant differences were observed in terms of distress (HADS, p = 0.297). Six (60%) patients continued with methylphenidate after study end. Main adverse events during study were palpitations and insomnia (30% of patients each). CONCLUSIONS: This pilot study suggests that methylphenidate may reduce asthenia and improve quality of life in breast cancer patients treated with docetaxel.

A genome-wide cell-free DNA methylation analysis identifies an episignature associated with metastatic luminal B breast cancer.

Breast cancers of the luminal B subtype are frequent tumors with high proliferation and poor prognosis. Epigenetic alterations have been found in breast tumors and in biological fluids. We aimed to profile the cell-free DNA (cfDNA) methylome of metastatic luminal B breast cancer (LBBC) patients using an epigenomic approach to discover potential noninvasive biomarkers. Plasma cfDNA was analyzed using the Infinium MethylationEpic array in a cohort of 14 women, including metastatic LBBC patients and nontumor controls. The methylation levels of cfDNA and tissue samples were validated with droplet digital PCR. The methylation and gene expression data of 582 primary luminal breast tumors and 79 nontumor tissues were obtained from The Cancer Genome Atlas (TCGA). We found an episignature of 1,467 differentially methylated CpGs that clearly identified patients with LBBC. Among the genes identified, the promoter hypermethylation of WNT1 was validated in cfDNA, showing an area under the ROC curve (AUC) of 0.86 for the noninvasive detection of metastatic LBBC. Both paired cfDNA and primary/metastatic breast tumor samples showed hypermethylation of WNT1. TCGA analysis revealed significant WNT1 hypermethylation in the primary tumors of luminal breast cancer patients, with a negative association between WNT1 methylation and gene expression. In this proof-of-principle study, we discovered an episignature associated with metastatic LBBC using a genome-wide cfDNA methylation approach. We also identified the promoter hypermethylation of WNT1 in cfDNA as a potential noninvasive biomarker for luminal breast cancer. Our results support the use of EPIC arrays to identify new epigenetic noninvasive biomarkers in breast cancer.