MicroRNA-based signatures impacting clinical course and biology of ovarian cancer: a miRNOmics study.

BACKGROUND: In Western countries, ovarian cancer (OC) still represents the leading cause of gynecological cancer-related deaths, despite the remarkable gains in therapeutical options. Novel biomarkers of early diagnosis, prognosis definition and prediction of treatment outcomes are of pivotal importance. Prior studies have shown the potentials of micro-ribonucleic acids (miRNAs) as biomarkers for OC and other cancers. METHODS: We focused on the prognostic and/or predictive potential of miRNAs in OC by conducting a comprehensive array profiling of miRNA expression levels in ovarian tissue samples from 17 non-neoplastic controls, and 60 tumor samples from OC patients treated at the Regina Elena National Cancer Institute (IRE). A set of 54 miRNAs with differential expression in tumor versus normal samples (T/N-deregulated) was identified in the IRE cohort and validated against data from the Cancer Genoma Atlas (TCGA) related to 563 OC patients and 8 non-neoplastic controls. The prognostic/predictive role of the selected 54 biomarkers was tested in reference to survival endpoints and platinum resistance (P-res). RESULTS: In the IRE cohort, downregulation of the 2 miRNA-signature including miR-99a-5p and miR-320a held a negative prognostic relevance, while upregulation of miR-224-5p was predictive of less favorable event free survival (EFS) and P-res. Data from the TCGA showed that downregulation of 5 miRNAs, i.e., miR-150, miR-30d, miR-342, miR-424, and miR-502, was associated with more favorable EFS and overall survival outcomes, while miR-200a upregulation was predictive of P-res. The 9 miRNAs globally identified were all included into a single biologic signature, which was tested in enrichment analysis using predicted/validated miRNA target genes, followed by network representation of the miRNA-mRNA interactions. CONCLUSIONS: Specific dysregulated microRNA sets in tumor tissue showed predictive/prognostic value in OC, and resulted in a promising biological signature for this disease.

Corrigendum to "Circulating Cancer Stem Cell-Derived Extracellular Vesicles as a Novel Biomarker for Clinical Outcome Evaluation".

[This corrects the article DOI: 10.1155/2019/5879616.].

Circulating Cancer Stem Cell-Derived Extracellular Vesicles as a Novel Biomarker for Clinical Outcome Evaluation.

The recent introduction of the "precision medicine" concept in oncology pushed cancer research to focus on dynamic measurable biomarkers able to predict responses to novel anticancer therapies in order to improve clinical outcomes. Recently, the involvement of extracellular vesicles (EVs) in cancer pathophysiology has been described, and given their release from all cell types under specific stimuli, EVs have also been proposed as potential biomarkers in cancer. Among the techniques used to study EVs, flow cytometry has a high clinical potential. Here, we have applied a recently developed and simplified flow cytometry method for circulating EV enumeration, subtyping, and isolation from a large cohort of metastatic and locally advanced nonhaematological cancer patients (N = 106); samples from gender- and age-matched healthy volunteers were also analysed. A large spectrum of cancer-related markers was used to analyse differences in terms of peripheral blood circulating EV phenotypes between patients and healthy volunteers, as well as their correlation to clinical outcomes. Finally, EVs from patients and controls were isolated by fluorescence-activated cell sorting, and their protein cargoes were analysed by proteomics. Results demonstrated that EV counts were significantly higher in cancer patients than in healthy volunteers, as previously reported. More interestingly, results also demonstrated that cancer patients presented higher concentrations of circulating CD31+ endothelial-derived and tumour cancer stem cell-derived CD133 + CD326- EVs, when compared to healthy volunteers. Furthermore, higher levels of CD133 + CD326- EVs showed a significant correlation with a poor overall survival. Additionally, proteomics analysis of EV cargoes demonstrated disparities in terms of protein content and function between circulating EVs in cancer patients and healthy controls. Overall, our data strongly suggest that blood circulating cancer stem cell-derived EVs may have a role as a diagnostic and prognostic biomarker in cancer.

Immunotherapy in HER2-positive breast cancer: state of the art and future perspectives.

Breast cancer (BC) is a complex disease with primary or acquired incurability characteristics in a significant part of patients. Immunotherapeutical agents represent an emerging option for breast cancer treatment, including the human epidermal growth factor 2 positive (HER2+) subtype. The immune system holds the ability to spontaneously implement a defensive response against HER2+ BC cells through complex mechanisms which can be exploited to modulate this response for obtaining a clinical benefit. Initial immune system modulating strategies consisted mostly in vaccine therapies, which are still being investigated and improved. However, the entrance of trastuzumab into the scenery of HER2+ BC treatment was the real game changing event, which embodied a dominant immune-mediated mechanism. More recently, the advent of the immune checkpoint inhibitors has caused a new paradigm shift for immuno-oncology, with promising initial results also for HER2+ BC. Breast cancer has been traditionally considered poorly immunogenic, being characterized by relatively low tumor mutation burden (TMB). Nevertheless, recent evidence has revealed high tumor infiltrating lymphocytes (TILs) and programmed cell death-ligand 1 (PD-L1) expression in a considerable proportion of HER2+ BC patients. This may translate into a higher potential to elicit anti-cancer response and, therefore, wider possibilities for the use and implementation of immunotherapy in this subset of BC patients. We are herein presenting and critically discussing the most representative evidence concerning immunotherapy in HER2+ BC cancer, both singularly and in combination with therapeutic agents acting throughout HER2-block, immune checkpoint inhibition and anti-cancer vaccines. The reader will be also provided with hints concerning potential future projection of the most promising immutherapeutic agents and approaches for the disease of interest.

Safety analysis, association with response and previous treatments of everolimus and exemestane in 181 metastatic breast cancer patients: A multicenter Italian experience.

PURPOSE: The everolimus and exemestane combination represents a treatment option for the endocrine sensitive metastatic breast cancer (MBC) patients. The toxicity profile reported in the Bolero 2 trial showed the feasibility in the selected patients. Few data are available for the unselected population. METHODS: In order to evaluate the safety in the unselected population of the clinical practice and to evaluate a possible association of toxicities with previous treatments, clinical data from 181 consecutive patients were retrospectively collected. RESULTS: Due to toxic events, everolimus dosage was reduced to 5 mg in 27% of patients. No association was found in the analysis between toxicity and number of prior therapies, neither between toxicity and response. In the multivariate analysis the previous exposure to anthracyclines for advanced disease represents the only predictive factor of developing grade ≥2 toxicity (OR = 2.85 CI 95% 1.07-7.59, p = 0.036). CONCLUSIONS: The association of everolimus and exemestane has confirmed to be a safe and effective treatment for endocrine sensitive MBC patients even in routine clinical practice. The rate of treatment discontinuation due to toxicity is low and none association between previous number of treatments and response or between toxicity and response was found.

Targeting immune response with therapeutic vaccines in premalignant lesions and cervical cancer: hope or reality from clinical studies.

Human papillomavirus (HPV) is widely known as a cause of cervical cancer (CC) and cervical intraepithelial neoplasia (CIN). HPVs related to cancer express two main oncogenes, i.e. E6 and E7, considered as tumorigenic genes; their integration into the host genome results in the abnormal regulation of cell cycle control. Due to their peculiarities, these oncogenes represent an excellent target for cancer immunotherapy. In this work the authors highlight the potential use of therapeutic vaccines as safe and effective pharmacological tools in cervical disease, focusing on vaccines that have reached the clinical trial phase. Many therapeutic HPV vaccines have been tested in clinical trials with promising results. Adoptive T-cell therapy showed clinical activity in a phase II trial involving advanced CC patients. A phase II randomized trial showed clinical activity of a nucleic acid-based vaccine in HPV16 or HPV18 positive CIN. Several trials involving peptide-protein-based vaccines and live-vector based vaccines demonstrated that these approaches are effective in CIN as well as in advanced CC patients. HPV therapeutic vaccines must be regarded as a therapeutic option in cervical disease. The synergic combination of HPV therapeutic vaccines with radiotherapy, chemotherapy, immunomodulators or immune checkpoint inhibitors opens a new and interesting scenario in this disease.

Origin of the pre-peak features in the oxygen K-edge x-ray absorption spectra of LaFeO3 and LaMnO3 studied by Ga substitution of the transition metal ion.

We report on experimental oxygen K-edge x-ray absorption near edge structure (XANES) spectra of the LaFe(1 - x)Ga(x)O(3) and LaMn(1 - x)Ga(x)O(3) series. Transition metal substitution by the 3d full shell Ga atom is mainly reflected in a systematic decrease of the pre-edge structures in the XANES spectra of the two series. This result shows that the associated states originate from the hybridization of oxygen 2p and unoccupied Fe (or Mn) 3d states. In order to gain insight into the states associated with the pre-edge spectral features, we have performed ab initio theoretical calculations based on multiple scattering theory. Simulations with variable cluster size and composition around the absorber oxygen in the LaFeO(3) and LaMnO(3) crystal structures were carried out. We find that the low-energy pre-peak is reproduced once the absorbing oxygen and the two nearest neighbour Fe (or Mn) ions are considered in the cluster. Conversely, higher energy pre-peaks only arise when the full oxygen coordination geometry around the two metal sites is taken into account, implying that their energy distance is a reflection of the strength of the oxygen ligand field. Substitutions of the two nearest neighbours by Ga atoms in the cluster of calculation lead to changes in the theoretical spectra that reasonably agree with the evolution of the pre-peaks in the experimental XANES spectra of both the series.

Unknown primary tumors.

An unknown primary tumor (UPT) is defined by the presence of a metastatic cancer without a known primary site of origin despite a standardized diagnostic workup. Clinically, UPTs show rapid progression and early dissemination, with signs and symptoms related to the metastatic site. The molecular bases of their biology remain largely unknown, with no evidence as to whether they represent a distinct biological entity. Immunohistochemistry remain the best diagnostic tool in term of cost-effectiveness, but the time-consuming "algorithmic process" it relies on has led to the application of new molecular techniques for the identification of the primary site of UPTs. For example, several microarray or miRNA classifications of UPTs have been used, with an accuracy in the prediction of the primary site as high as 90%. It should be noted that validating a prediction of tissue origin is challenging in these patients, since most of them will never have a primary site identified. Moreover, prospective studies to determine whether selection of treatment options based on such profiling methods actually improves patient outcome are still missing. In the last few years functional imaging (i.e. FDG-PET/CT) has gained a main role in the detection of the site of origin of UPTs and is currently recommended by the European Association of Nuclear Medicine. However, despite recent refinements in the diagnostic workup, the site of origin of UPT often remains elusive. As a consequence, treatment of patients with UPT is still empirical and inadequate.

Tyrosine kinase inhibitors.

Over the last ten years, several new and therapeutically relevant cancer drugs targeting tyrosine kinases signaling pathways have been developed. Tyrosine kinase inhibitors (TKIs) are a pharmaceutical class of small molecules, orally available, well-tolerated, worldwide approved drugs for the treatment of several neoplasms, including lung, breast, kidney and pancreatic cancer as well as gastro-intestinal stromal tumors and chronic myeloid leukemia. This comprehensive review focuses on the most relevant members of the first and the second generation TKIs designed to interact with receptor and nonreceptor TKs. Attention is mainly focused on molecular mechanisms in in vitro and in vivo models related to the clinical activity of the drugs and to the development of resistance to treatment, still the major challenge in cancer research and care.

Antiferromagnetic-paramagnetic insulating transition in Cr-doped V(2)O(3) investigated by EXAFS analysis.

We have performed extended x-ray absorption fine-structure (EXAFS) spectroscopy on a 2.8% Cr-doped V(2)O(3) sample, with the aim of studying its structural evolution in a wide temperature range across the paramagnetic-antiferromagnetic insulating phase transition at T(c). The data were registered with two different set-ups in fluorescence and transmission geometries, for polarized and unpolarized spectra, respectively. Our idea, based on previous experiments reported in the literature, is that extended structural modifications of the nominal trigonal symmetry are present in the paramagnetic insulating phase for several tens of degrees above T(c), involving further-nearest-neighbor vanadium ions. Our data confirm that the paramagnetic insulating phase is not structurally homogeneous in a temperature range of about 30 K around T(c), where local distortions of monoclinic symmetry involving further-nearest neighbors are present. Moreover, the analysis of the absorption profile at Cr K-edge suggests that Cr ions enter the lattice randomly. We finally analyze our findings in light of current theoretical models.

Dynamics of cell cycle phase perturbations by trabectedin (ET-743) in nucleotide excision repair (NER)-deficient and NER-proficient cells, unravelled by a novel mathematical simulation approach.

OBJECTIVES: Trabectedin (ET-743, Yondelis) is a natural marine product, with antitumour activity, currently in phase II/III clinical trials. Previous studies have shown that cells hypersensitive to ultraviolet (UV)-rays because of nucleotide excision repair (NER) deficiency, were resistant to trabectedin. The purpose of this study was to investigate whether this resistance was associated with different drug-induced cell cycle perturbations. MATERIALS AND METHODS: An isogenic NER-proficient cellular system (CHO-AA8) and a NER-deficient one (CHO-UV-96), lacking functional ERCC-1, were studied. Flow cytometric assays showed progressive accumulation of cells in G2 + M phase in NER-proficient but not in NER-deficient cells. Applying a computer simulation method, we realized that the dynamics of the cell cycle perturbations in all phases were complex. RESULTS: Cells exposed to trabectedin during G1 and G2 + M first experienced a G1 block, while those exposed in S phase were delayed in S and G2 + M phases but eventually divided. In the presence of functional NER, exit from the G1 block was faster; then, cells progressed slowly through S phase and were subsequently blocked in G2 + M phase. This G2 + M processing of trabectedin-induced damage in NER-proficient cells was unable to restore cell cycling, suggesting a difficulty in repairing the damage. CONCLUSIONS: This might be due either to important damage left unrepaired by previous G1 repair, or that NER activity itself caused DNA damage, or both. We speculate that in UV-96 cells repair mechanisms other than NER are activated both in G1 and G2 + M phases.

A phase II study of dose-dense epirubicin plus cyclophosphamide followed by docetaxel plus capecitabine and pegfilgrastim support as preoperative therapy for patients with stage II, IIIA breast cancer.

BACKGROUND: This phase II study was conducted to evaluate tumor response rate and safety profile of dose-dense epirubicin plus cyclophosphamide followed by docetaxel plus capecitabine given preoperatively to patients with stage II or IIIA breast cancer. PATIENTS AND METHODS: Patients underwent four cycles of dose-dense cyclophosphamide 600 mg/m(2) and epirubicin 90 mg/m(2) every 2 weeks followed by two cycles of docetaxel 36 mg/m(2) on days 1, 8, and 15 plus capecitabine 1250 mg/m(2) on days 5-18 every 4 weeks, with prophylactic pegfilgrastim. The primary objective of the study was to determine the incidence of pathologic complete response defined as the absence of invasive or in situ cancer in the breast and the axillary nodes at definitive surgery. RESULTS: Forty-four patients were enrolled in the study and 41 (93%) were assessable for response to chemotherapy. An objective clinical response was observed in 38 (93%) patients. Seven patients (17.1%) exhibited a pathologic complete response. Breast-conserving surgery was carried out in 36 (88%) patients. Grade 3/4 neutropenia occurred in 4.3% of 252 administered chemotherapy cycles. No febrile neutropenia, cardiac toxicity, thrombocytopenia or other serious adverse event was registered. CONCLUSION: The sequential combination of dose-dense epirubicin plus cyclophosphamide followed by docetaxel plus capecitabine is an effective and well-tolerated neo-adjuvant chemotherapy for stage II, IIIA breast cancer.

Timed flat infusion of 5-fluorouracil increases the tolerability of 5-fluorouracil/docetaxel regimen in metastatic breast cancer: a dose-finding study.

A dose-finding study was undertaken to determine the maximum-tolerated dose, and the recommended dose of docetaxel in combination with 12-h timed (22:00-10:00) flat infusion of 5-fluorouracil (5-FU) in metastatic breast cancer patients. This schedule seems to reduce the occurrence of stomatitis of the docetaxel and infusional 5-FU regimen.

X-ray absorption spectroscopy: state-of-the-art analysis.

State-of-the-art techniques for analysing X-ray absorption spectra are reviewed, with an eye to biological applications. Recent attempts to perform full spectral fitting of the XANES energy region and beyond for the purpose of structural analysis have met with encouraging success. The present paper analyses the theoretical motivations behind this success and indicates routes for future improvements. The theoretical background is not entirely new, although the point of view is, and some sections and appendices present material that the Authors believe has never been published before. The aim of this paper is to provide a theoretical analysis that is as self-contained as possible.

The MXAN procedure: a new method for analysing the XANES spectra of metalloproteins to obtain structural quantitative information.

The first quantitative analyses are reported of the Fe K-edge polarized X-ray absorption near-edge structure (XANES) of a single crystal of the iron protein carbonmonoxy-myoglobin (MbCO) and of its cryogenic photoproduct Mb*CO. The CO-Fe-heme local structure has been determined using a novel fitting procedure, named MXAN, which is able to fit the XANES part (from the edge to about 200 eV) of experimental X-ray absorption data. This method is based on the comparison between the experimental spectrum and several theoretical spectra that are generated by changing the relevant geometrical parameters of the site around the absorbing atom. The theoretical spectra are derived in the framework of the full multiple-scattering approach. The MXAN procedure is able to recover information about the symmetry and atomic distances, and the solution is found to be independent of the starting conditions. The extracted local structure of Mb*CO includes an Fe-CO distance of 3.08 (7) A, with a tilting angle between the heme normal and the Fe-C vector of 37 (7) degrees and a bending angle between the Fe-C vector and the C-O bond of 31 (5) degrees

Local chiral-symmetry breaking in globally centrosymmetric crystals.

A thorough tensor analysis of the Bragg-forbidden reflection (00.3)(h) in corundum systems having a global center of inversion, such as V2O3 and alpha-Fe2O3, shows that anomalous x-ray resonant diffraction can access chiral properties related to the dipole-quadrupole (E1-E2) channel via an interference with the pure quadrupole-quadrupole (E2-E2) process. This is also confirmed by independent ab initio numerical simulations. In such a way, it becomes possible to detect chiral quantities in systems where dichroic absorption techniques are ineffective.

Expression of glycoprotein 90K in human malignant pleural mesothelioma: correlation with patient survival.

The expression of the tumour-associated glycoprotein 90K in patients with malignant pleural mesothelioma (MM) has not been described. This study used enzyme-linked immunoassay (ELISA) to measure 90K in pleural effusions (PEs) and sera from patients with MM (n=28), lung cancer (LC) (n=14) and benign pleural disease (BPD) (n=15). Immunohistochemistry was used to investigate 90K expression in MM and LC tissue sections. The expression of 90K was further evaluated in vitro by ELISA and western blot analysis of conditioned media and cellular extracts of MM, LC and normal human mesothelial (NHM) cell cultures. Finally, the relationships between 90K expression in MM and patient age and survival were studied. The mean 90K level was significantly higher (p<0.05) in PEs of MM patients (11.0+/-6.6 microg/ml) than in LC (6.1+/-3.2 microg/ml) or BPD (6.2+/-5.0 microg/ml) patients. Immunohistochemistry showed a positive reaction for 90K in MM biopsy sections and positive staining limited to inflammatory infiltrates in LC sections. The level of 90K was significantly higher in cell culture media of MM than of LC or NHM (p<0.001). Bands representing proteins with molecular weight of approximately 90 kDa were detected by western blot in MM cellular extracts. An inverse correlation between PE 90K levels and MM patient age (r=-0.45; p=0.017) and a positive correlation between serum 90K levels and MM patient survival (r=0.62; p=0.006) were detected by linear regression analysis. Kaplan-Meier univariate analysis showed increased survival probability for MM patients with serum 90K level >7.3 microg/ml (log rank, p<0.05). This is the first report in MM of the expression of 90K and of its potential diagnostic and prognostic application.

A multiple-scattering theory of circular and linear dichroism for photoemission and photoabsorption.

A unified treatment of circular dichroism, both natural and magnetic, in (spin-resolved) photoemission and photoabsorption from core levels is given, valid in the many-body case and for extended systems, together with an extension of the formalism to treat linear dichroism. The reduction of this scheme to a one-electron picture in the framework of multiple scattering theory is briefly discussed and shows the intimate connection of the two spectroscopies via a generalized optical theorem. Plausibility arguments are given that in correlated d-band systems screening and relaxation effects are not so drastic as in other cases, due to the autoscreening action of the excited photoelectron, so that the final density of states is much like the initial unperturbed one. It is shown how to exploit this point of view to obtain in favorables cases separated orbital and spin moment radial (surface) distribution maps from dichroic magnetic EXAFS spectra (photoelectron diffraction patterns) related to the ground state. Dichroic natural spectra, both in photoemission and absorption, are shown to be sensitive only to atoms in chiral geometry.

Symmetry role on the preedge X-ray absorption fine structure at the metal K edge.

The preedge features in a system with "even" symmetry, apart from quadrupolar transition contribution, are mainly dipolar in character, associated with the existence of unoccupied states made up of mixed cation-4p with higher-neighboring cation-3d orbitals, and reflect the density of states due to the medium-range order of the system. In "odd" symmetry materials these preedge features are the result of a transition from the 1s to a final density of states of p symmetry due to an unsymmetrical mixing of the ligand wave functions with the central cation 3d orbitals, similar to atetrahedral configuration. These results are validated for Fe as a photoabsorber by comparing XAS spectra of Fe2SiO4 (fayalite) to ab initio full multiple scattering calculations at the Fe K edge, but pertain to all systems containing sixfold-coordinated cations.

Adhesion to 90K (Mac-2 BP) as a mechanism for lymphoma drug resistance in vivo.

A cell-adhesive protein of the human serum, 90K binds galactin-3, beta1-integrins, collagens, and fibronectin, and it is of importance in cell-cell and cell-extracellular matrix adhesion. Serum 90K levels in 137 patients with lymphoma were measured by enzyme-linked immunosorbent assay. Compared with healthy controls, pretreatment serum 90K levels in patients with lymphoma were elevated (P <.001). Of 97 patients who showed objective response to treatment, 20 (21%) had pretreatment 90K levels above the normal cutoff compared with 17 (53%) of 32 patients who did not respond (P =.002). When used as a plastic-immobilized substrate, 90K caused a significant reduction in chemotherapy-induced apoptosis of Jurkat T lymphoma cells. This finding could explain the lack of response in lymphoma patients with high 90K serum levels.