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Abstract Despite progress in breast cancer treatment, a significant portion of patients still relapse because of drug resistance. The involvement of microRNAs in cancer progression and chemotherapy response is well established. Therefore, this study aimed to elucidate the dysregulation of the microRNA-449 family (specifically, microRNA-449a, microRNA-449b-5p, and microRNA-449c-5p) and its impact on resistance to doxorubicin, a commonly used chemotherapeutic drug for the treatment of triple-negative breast cancer. We found that the microRNA-449 family is downregulated in triple-negative breast cancer and demonstrated its potential as a diagnostic biomarker. Besides, our findings indicate that the downregulation of the microRNA-449 family is mediated by the microRNAs-449/SIRT1-HDAC1 negative feedback loop. Moreover, it was found that the microRNA-449 family dysregulates the fatty acid metabolism by targeting ACSL4, which is a potential prognostic biomarker that mediates doxorubicin response through regulation of the drug extrusion pump ABCG2. Altogether, our results suggest that the microRNA-449 family might be a potential therapeutic target for the treatment of triple-negative breast cancer since it is implicated in doxorubicin response through ACSL4/ABCG2 axis regulation. Ultimately, our results also highlight the value of microRNAs-449 and ACSL4 as diagnostic and prognostic biomarkers in triple-negative breast cancer. Proposed model of miRNAs-449 downregulation in TNBC and doxorubicin response. MiRNAs-449 are downregulated in TNBC through a negative feedback loop with SIRT1 and HDAC1. Moreover, ACSL4 increases ABCG2 expression, thus diminishing the intracellular doxorubicin concentration and promoting doxorubicin resistance. MiRNAs-449 overexpression downregulates the ACSL4/ABCG2 axis and sensitizes doxorubicin-resistant cells to doxorubicin. Created with BioRender. TNBC: triple-negative breast cancer; DOX: doxorubicin; SIRT1: Sirtuin 1; HDAC1: Histone deacetylase 1; ACSL4: Acyl-CoA Synthetase Long-Chain Family Member 4; ABCG2: ATP-binding cassette superfamily G member 2.

A substantial proportion of HER2+ breast cancer patients do not benefit from HER2-targeted therapy. Here, the authors identify a population of cancer-associated fibroblasts involved in the suppression of trastuzumab-induced ADCC that can be pharmacologically targeted to raise treatment effectiveness in unresponsive tumors.

BackgroundThe SOLTI-1301 AGATA study aimed to assess the feasibility of a multi-institutional molecular screening program to better characterize the genomic landscape of advanced breast cancer (ABC) and to facilitate patient access to matched-targeted therapies in Spain.MethodsDNA sequencing of 74 cancer-related genes was performed using FFPE tumor samples in three different laboratories with three different gene panels. A multidisciplinary advisory board prospectively recommended potential targeted treatments. The primary objective was to determine the success of matching somatic DNA alteration to an experimental drug/drug class.ResultsBetween September 2014 and July 2017, 305 patients with ABC from 10 institutions were enrolled. Tumor sequencing was successful in 260 (85.3%) patients. Median age was 54 (29-80); most tumors were hormone receptor-positive/HER2-negative (74%), followed by triple-negative (14.5%) and HER2-positive (11.5%). Ninety-seven (37%) tumor samples analyzed proceeded from metastatic sites. Somatic mutations were identified in 163 (62.7%) patients, mostly in PIK3CA (34%), TP53 (22%), AKT1 (5%), ESR1 (3%), and ERBB2 (3%) genes. Significant enrichment of AKT1 mutation was observed in metastatic versus primary samples (9% vs. 2%; p=0.01). Genome-driven cancer therapy was recommended in 45% (n=116) of successfully screened patients, 11% (n=13) of whom finally received it. Among these patients, 46.2% had a PFS of ≥6 months on matched therapy.ConclusionsAGATA is the first nationwide molecular screening program carried out in Spain and we proved that implementing molecular data in the management of ABC is feasible. Although these results are promising, only 11% of the patients with genome-driven cancer therapy received it.

Abstract Background Pathological complete response (pCR) after neoadjuvant chemotherapy (NAC) in triple-negative breast cancer (TNBC) varies between 30 and 40% approximately. To provide further insight into the prediction of pCR, we evaluated the role of an epigenetic methylation-based signature. Methods Epigenetic assessment of DNA extracted from biopsy archived samples previous to NAC from TNBC patients was performed. Patients included were categorized according to previous response to NAC in responder (pCR or residual cancer burden, RCB = 0) or non-responder (non-pCR or RCB > 0) patients. A methyloma study was performed in a discovery cohort by the Infinium HumanMethylation450 BeadChip (450K array) from Illumina. The epigenetic silencing of those methylated genes in the discovery cohort were validated by bisulfite pyrosequencing (PyroMark Q96 System version 2.0.6, Qiagen) and qRT-PCR in an independent cohort of TN patients and in TN cell lines. Results Twenty-four and 30 patients were included in the discovery and validation cohorts, respectively. In the discovery cohort, nine genes were differentially methylated: six presented higher methylation in non-responder patients (LOC641519, LEF1, HOXA5, EVC2, TLX3, CDKL2) and three greater methylation in responder patients (FERD3L, CHL1, and TRIP10). After validation, a two-gene (FER3L and TRIP10) epigenetic score predicted RCB = 0 with an area under the ROC curve (AUC) = 0.905 (95% CI = 0.805–1.000). Patients with a positive epigenetic two-gene score showed 78.6% RCB = 0 versus only 10.7% RCB = 0 if signature were negative. Conclusions These results suggest that pCR in TNBC could be accurately predicted with an epigenetic signature of FERD3L and TRIP10 genes. Further prospective validation of these findings is warranted.

Downregulation of miR-33b has been documented in many types of cancers and is being involved in proliferation, migration, and epithelial–mesenchymal transition (EMT). Furthermore, the enhancer of zeste homolog 2-gene (EZH2) is a master regulator of controlling the stem cell differentiation and the cell proliferation processes. We aim to evaluate the implication of miR-33b in the EMT pathway in HER2+ breast cancer (BC) and to analyze the role of EZH2 in this process as well as the interaction between them. miR-33b is downregulated in HER2+ BC cells vs healthy controls, where EZH2 has an opposite expression in vitro and in patients’ samples. The upregulation of miR-33b suppressed proliferation, induced apoptosis, reduced invasion, migration and regulated EMT by an increase of E-cadherin and a decrease of ß-catenin and vimentin. The silencing of EZH2 mimicked the impact of miR-33b overexpression. Furthermore, the inhibition of miR-33b induces cell proliferation, invasion, migration, EMT, and EZH2 expression in non-tumorigenic cells. Importantly, the Kaplan–Meier analysis showed a significant association between high miR-33b expression and better overall survival. These results suggest miR-33b as a suppressive miRNA that could inhibit tumor metastasis and invasion in HER2+ BC partly by impeding EMT through the repression of the MYC–EZH2 loop.

The receptor activator of nuclear factor kappa B (RANK) is becoming recognized as a master regulator of tumorigenesis, yet its role in gynecological cancers remains mostly unexplored. We investigated whether there is a gradation of RANK protein and mRNA expression in epithelial ovarian cancer (EOC) according to malignancy and tumor staging. Immunohistochemical expression of RANK was examined in a cohort of 135 (benign n = 29, borderline n= 23 and malignant n = 83) EOCs. Wild type and truncated RANK mRNA isoform quantification was performed in a cohort of 168 (benign n = 26, borderline n = 13 and malignant n = 129) EOCs. RANK protein and mRNA values were increased in malignant vs. benign or borderline conditions across serous, mucinous and endometrioid cancer subtypes. Additionally, a trend of increased RANK values with staging was observed for the mucinous and serous histotype. Thus, increased expression of RANK appears associated with the evolution of disease to the onset of malignancy in EOC. Moreover, in some EOC histotypes, RANK expression is additionally associated with clinicopathological markers of tumor aggressiveness, suggesting a role in further progression of tumor activity.

Purpose: Breast cancer (BC) in very young women (BCVY) is more aggressive than in older women. The purpose of this study was to evaluate the relevance of a range of clinico-pathological factors in the prognosis of BCVY patients. Methods: We retrospectively analyzed 258 patients diagnosed with BCVY at our hospital from 1998 to 2014; the control group comprised 101 older patients with BC. We correlated clinicopathological factors, treatments, relapse and exitus with age and with previously published miRNA expression data. Results: We identified some significant differences in risk factors between BCVY and older patients. The age at menarche, number of pregnancies, and age at first pregnancy were lower in the BCVY group and had a greater probability of recurrence and death in all cases. Lymph node-positive patients in the BCVY group are associated with a worse prognosis ( P = .02), an immunohistochemical HER2 + subtype, and disease relapse ( P = .03). Moreover, there was a shorter time between diagnosis and first relapse in BCVY patients compared with controls, and they were more likely to die from the disease ( P = .002). Finally, from our panel of miRNAs deregulated in BC, reduced miR-30c expression was associated with more aggressive BC in very young patients, lower overall survival, and with axillary lymph node metastases. Conclusions: Patient age and axillary lymph node status post-surgery are independent and significant predictors of distant disease-free survival, local recurrence-free survival, and overall survival. The HER2 + subtype and lower miR-30c expression are related to poor prognosis in lymph node-positive young BC patients.

The system integrated by the receptor activator of nuclear factor kappa B (RANK) and its ligand, RANKL, modulates the role of hormones in the genesis and progression of breast tumors. We investigated whether the expression of RANK was related with clinicopathological features of primary endometrial tumors. Immunohistochemistry was used in an endometrial cancer tissue array containing samples from 36 tumors. The amount of RANK mRNA was examined in a tissue scan cDNA array containing cDNA from 40 tumors. Normal endometrium was examined for comparison. Immunohistochemical analyses showed that RANK expression was higher in malignant than in normal endometrium (p < 0.05). RANK expression was related to histological grade (Pearson correlation index = 0.484, p < 0.001), but not to tumor stage or to age of the women. The gene expression was similar in malignant and normal endometrium. The study of RANK isoforms confirmed that the overall relative abundance of the three clearly identified transcripts was similar in normal and pathological endometrium. RANK protein expression increased from normal to malignant endometrium, and the expression level was related with tumor grade but not with stage or the age of subjects in endometrial cancer. In contrast, similar comparisons showed no change in RANK gene expression.

Introduction: Strongyloides has been shown to infrequently mimic inflammatory bowel disease (IBD) or to disseminate when a patient with IBD and unrecognized strongyloides is treated with immunosupression. Case report: A man from Ecuador, living in Spain for years, with a history of type 2 diabetes mellitus and psoriasis treated with topical corticosteroids, was admitted to the hospital with an 8-month history of diarrhoea. Blood tests showed hyperglycemia, hyponatremia, elevated CRP and faecal calprotectin. Colonoscopy suggested IBD. The patient improved with steroids, pending biopsy results, and he was discharged. Biopies were compatible with IBD, but careful examination revealed strongyloides. He was given a prescription of albendazole. He had to be readmitted due to SIADH, which resolved with fluid restriction. Upon discharge albendazole was prescribed again. The patient skipped most of the out-patient-clinic visits. He returned a year later on 10 mg/week methotrexate, asymptomatic, with 20% eosinophilia, and admitting he had never taken the strongyloides treatment for economical reasons. He then received a week of oral albendazol at the hospital. Biopsies and blood cell count were afterwards normal (eosinophils 3.1%) and serology for strongyloides antibodies was negative. Discussion: This case is of interest for four rarely concurring reasons. It's a worm infection that mimics IBD; the infection was diagnosed by colon biopsy; the infection caused a SIADH; and, most interestingly, even though the patient is on immunosupression, he remains asymptomatic.

Due to the lack of specific targets, cytotoxic chemotherapy still represents the common standard treatment for triple-negative breast patients. Despite the harmful effect of chemotherapy on tumor cells, there is evidence that treatment could modulate the tumor microenvironment in a way favoring the propagation of the tumor. In addition, the lymphangiogenesis process and its factors could be involved in this counter-therapeutic event. In our study, we have evaluated the expression of the main lymphangiogenic receptor VEGFR3 in two triple-negative breast cancer in vitro models, resistant or not to doxorubicin treatment. The expression of the receptor, at mRNA and protein levels, was higher in doxorubicin-resistant cells than in parental cells. In addition, we confirmed the upregulation of VEGFR3 levels after a short treatment with doxorubicin. Furthermore, VEGFR3 silencing reduced cell proliferation and migration capacities in both cell lines. Interestingly, high VEGFR3 expression was significantly positively correlated with worse survival in patients treated with chemotherapy. Furthermore, we have found that patients with high expression of VEGFR3 present shorter relapse-free survival than patients with low levels of the receptor. In conclusion, elevated VEGFR3 levels correlate with poor survival in patients and with reduced doxorubicin treatment efficacy in vitro. Our results suggest that the levels of this receptor could be a potential marker of meager doxorubicin response. Consequently, our results suggest that the combination of chemotherapy and VEGFR3 blockage could be a potentially useful therapeutic strategy for the treatment of triple-negative breast cancer.

Treatment for the HER2+ breast cancer subtype is still unsatisfactory, despite breakthroughs in research. The discovery of various new molecular mechanisms of transcription factors may help to make treatment regimens more effective. The transcription factor SALL4 has been related to aggressiveness and resistance therapy in cancer. Its molecular mechanisms and involvement in various signaling pathways are unknown in the HER2+ breast cancer subtype. In this study, we have evaluated the implication of SALL4 in the HER2+ subtype through its expression in patients’ samples and gain and loss of function in HER2+ cell lines. We found higher SALL4 expression in breast cancer tissues compared to healthy tissue. Interestingly, high SALL4 expression was associated with disease relapse and poor patient survival. In HER2+ cell lines, transient overexpression of SALL4 modulates PI3K/AKT signaling through regulating PTEN expression and BCL2, which increases cell survival and proliferation while reducing the efficacy of trastuzumab. SALL4 has also been observed to regulate the epithelial–mesenchymal transition and stemness features. SALL4 overexpression significantly reduced the epithelial markers E-cadherin, while it increased the mesenchymal markers β-catenin, vimentin and fibronectin. Furthermore, it has been also observed an increased expression of MYC, an essential transcription factor for regulating epithelial-mesenchymal transition and/or cancer stem cells. Our study demonstrates, for the first time, the importance of SALL4 in the HER2+ subtype and partial regulation of trastuzumab sensitivity. It provides a viable molecular mechanism-driven therapeutic strategy for an important subset of HER2-overexpressing patients whose malignancies are mediated by SALL4 expression.

Aurora kinases (AURK) are key regulators of the mitotic spindle formation. AURK is frequently overexpressed in ovarian cancer and this overexpression has been frequently associated with prognosis in these tumours. Interestingly, AURK have been shown to interact with DNA repair mechanisms and other cell cycle regulators. These functions have brought light to Aurora family as a potential target for anticancer therapy. In the last years, two clinical trials with different AURK inhibitors have shown activity in epithelial and clear-cell ovarian cancer. Although there is a lack of predictive factors of AURK inhibition activity, recent trials have identified some candidates. This review will focus in the functions of the AURK family, its role as prognostic factor in epithelial ovarian cancer and potential clinical implications.

A chromosomal region that includes the gene encoding HER2, a receptor tyrosine kinase (RTK), is amplified in 20% of breast cancers. Although these tumors tend to respond to drugs directed against HER2, they frequently become resistant and resume their malignant progression. Gene amplification in double minutes (DMs), which are extrachromosomal entities whose number can be dynamically regulated, has been suggested to facilitate the acquisition of resistance to therapies targeting RTKs. Here we show that ~30% of HER2-positive tumors show amplification in DMs. However, these tumors respond to trastuzumab in a similar fashion than those with amplification of the HER2 gene within chromosomes. Furthermore, in different models of resistance to anti-HER2 therapies, the number of DMs containing HER2 is maintained, even when the acquisition of resistance is concomitant with loss of HER2 protein expression. Thus, both clinical and preclinical data show that, despite expectations, loss of HER2 protein expression due to loss of DMs containing HER2 is not a likely mechanism of resistance to anti-HER2 therapies.