126 results on '"Signore, Michele"'
Search Results
102. Activity of the BH3 Mimetic ABT-737 on Polycythemia Vera (PV) Erythroid Precursor Cells
- Author
-
Tafuri, Agostino, primary, Pedini, Francesca, primary, Francescangeli, Federica, primary, Signore, Michele, primary, Foa, Robert, primary, Girelli, Gabriella, primary, Ruggero, De Maria, primary, and Ann, Zeuner, primary
- Published
- 2008
- Full Text
- View/download PDF
103. Quantum dots for biomedical applications
- Author
-
Signore, Michele, primary, Liotta, Lance A, additional, Maria, Ruggero De, additional, and Petricoin, Emanuel F, additional
- Published
- 2008
- Full Text
- View/download PDF
104. Reverse-phase protein microarrays: application to biomarker discovery and translational medicine
- Author
-
VanMeter, Amy, primary, Signore, Michele, additional, Pierobon, Mariaelena, additional, Espina, Virginia, additional, Liotta, Lance A, additional, and Petricoin III, Emanuel F, additional
- Published
- 2007
- Full Text
- View/download PDF
105. Chemotherapy-Induced Thrombocytopenia Derives from the Selective Death of Megakaryocyte Progenitors and Can Be Rescued by Stem Cell Factor
- Author
-
Zeuner, Ann, primary, Signore, Michele, additional, Martinetti, Daniela, additional, Bartucci, Monica, additional, Peschle, Cesare, additional, and De Maria, Ruggero, additional
- Published
- 2007
- Full Text
- View/download PDF
106. Activity of the BH3 Mimetic ABT-737 on Polycythemia Vera (PV) Erythroid Precursor Cells
- Author
-
agostino tafuri, Pedini, Francesca, Francescangeli, Federica, Signore, Michele, Foa, Robert, Girelli, Gabriella, Ruggero, Maria, and Ann, Zeuner
- Subjects
hemic and lymphatic diseases ,Immunology ,Cell Biology ,Hematology ,Biochemistry - Abstract
Malignant hemopoietic cells are often characterized by ineffective death pathways, resulting in enhanced resistance to apoptosis and ultimately in the survival/expansion of the abnormal clone. Mechanistic studies have been undertaken to identify the aberrant signal transduction pathway and to develop small-molecule inhibitors targeting deregulated modules. Regulators of apoptotic pathways play a key role in the control of erythroid cell expansion; in particularl Bcl-XL is essential for erythroid cell development and, together with Bcl-2, it protects erythroblast survival from cytotoxic stimuli. Previous studies on erythroid cells derived from polycythemia vera (PV) patients have revealed an increased expression of Bcl-XL associated with cell survival in the absence of erythropoietin. The discovery of the JAK2V617F mutation in the vast majority of PV patients and its association with an increased resistance to apoptosis induced by death receptors prompted us to investigate the correlation between JAK2V617F and the expression of anti-apoptotic Bcl-2 family members and to explore the activity of a Bcl-2 inhibitor on PV erythroid cells. ABT-737 is a synthetic small-molecule inhibitor that binds with high affinity to Bcl-2 and Bcl-XL, poorly to Mcl-1, promoting apoptosis as single agent in malignant hemopoietic cells. In this study we analyzed pure populations of primary erythroid precursors obtained from CD34+ cells of healthy donors and PV patients to investigate the expression of Bcl-2, Bcl-XL and Mcl-1. We found that the expression of Bcl-XL and Bcl-2 was increased in PV patients compared to controls, while Mcl-1 levels did not significantly differ. Then we analyzed Bcl-2 and Bcl-XL expression in erythroblasts derived from PV patients clustered on the basis of a different JAK2V617F allele burden. We found that on average the expression of both Bcl-2 and Bcl-XL was comparable to controls in PV erythroblasts with low/null mutation rates while both proteins were significantly (P< 0.001 and P
- Published
- 2008
107. Increased death receptor resistance and FLIPshort expression in polycythemia vera erythroid precursor cells
- Author
-
Zeuner, Ann, primary, Pedini, Francesca, additional, Signore, Michele, additional, Ruscio, Giusy, additional, Messina, Carlo, additional, Tafuri, Agostino, additional, Girelli, Gabriella, additional, Peschle, Cesare, additional, and De Maria, Ruggero, additional
- Published
- 2006
- Full Text
- View/download PDF
108. Inhibition of DNA Methylation Sensitizes Glioblastoma for Tumor Necrosis Factor–Related Apoptosis-Inducing Ligand–Mediated Destruction
- Author
-
Eramo, Adriana, primary, Pallini, Roberto, additional, Lotti, Fiorenza, additional, Sette, Giovanni, additional, Patti, Mariella, additional, Bartucci, Monica, additional, Ricci-Vitiani, Lucia, additional, Signore, Michele, additional, Stassi, Giorgio, additional, Larocca, Luigi M., additional, Crinò, Lucio, additional, Peschle, Cesare, additional, and De Maria, Ruggero, additional
- Published
- 2005
- Full Text
- View/download PDF
109. Oxidative stress and mitochondrial glutathione in human lymphocytes exposed to clinically relevant anesthetic drug concentrations
- Author
-
Delogu, Giovanna, primary, Antonucci, Adriana, additional, Moretti, Sonia, additional, Marandola, Maurizio, additional, Tellan, Guglielmo, additional, Signore, Michele, additional, and Famularo, Giuseppe, additional
- Published
- 2004
- Full Text
- View/download PDF
110. Antibody Validation by Western Blotting.
- Author
-
Signore, Michele and Reeder, K. Alex
- Published
- 2012
- Full Text
- View/download PDF
111. Stem cell factor protects erythroid precursor cells from chemotherapeutic agents via up-regulation of BCL-2 family proteins
- Author
-
Zeuner, Ann, primary, Pedini, Francesca, additional, Signore, Michele, additional, Testa, Ugo, additional, Pelosi, Elvira, additional, Peschle, Cesare, additional, and De Maria, Ruggero, additional
- Published
- 2003
- Full Text
- View/download PDF
112. Heat shock proteins and their role in heart injury
- Author
-
Delogu, Giovanna, primary, Signore, Michele, additional, Mechelli, Andrea, additional, and Famularo, Giuseppe, additional
- Published
- 2002
- Full Text
- View/download PDF
113. A new bioavailable fenretinide formulation with antiproliferative, antimetabolic, and cytotoxic effects on solid tumors
- Author
-
Angelita Costantino, Angelo Peschiaroli, Gerry Melino, Marco Tartaglia, Maria Laura De Angelis, Lucilla Bongiorno-Borbone, Marta Baiocchi, Alessandro Giuliani, Michele Signore, Alessandra Boe, Adriana Eramo, Ruggero De Maria, Alessandro Bruselles, Ann Zeuner, Paola Contavalli, Isabella Orienti, Federica Francescangeli, Toshio Kitamura, Massimo Spada, Filippo La Torre, Valentina Salvati, Giovanni Sette, Toshihiko Oki, Lello Zolla, Katia Fecchi, Signore, Michele [0000-0002-0262-842X], Melino, Gerry [0000-0001-9428-5972], Zeuner, Ann [0000-0002-8295-3715], Apollo - University of Cambridge Repository, Orienti I., Francescangeli F., De Angelis M.L., Fecchi K., Bongiorno-Borbone L., Signore M., Peschiaroli A., Boe A., Bruselles A., Costantino A., Eramo A., Salvati V., Sette G., Contavalli P., Zolla L., Oki T., Kitamura T., Spada M., Giuliani A., Baiocchi M., La Torre F., Melino G., Tartaglia M., De Maria R., and Zeuner A.
- Subjects
0301 basic medicine ,Cancer Research ,Lung Neoplasms ,Fenretinide ,DNA damage ,Colorectal cancer ,Immunology ,Antineoplastic Agents ,Apoptosis ,fenretinide, cyclodextrin, bioavailable formulation, solid tumors ,p38 Mitogen-Activated Protein Kinases ,Article ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,chemistry.chemical_compound ,Mice ,0302 clinical medicine ,Settore MED/04 - PATOLOGIA GENERALE ,Cell Line, Tumor ,medicine ,Tumor Cells, Cultured ,Animals ,Humans ,lcsh:QH573-671 ,Settore BIO/10 ,PI3K/AKT/mTOR pathway ,Cell Proliferation ,lcsh:Cytology ,Cancer stem cells ,Cell Cycle ,Cell Biology ,medicine.disease ,Xenograft Model Antitumor Assays ,Bioavailability ,030104 developmental biology ,chemistry ,Cell culture ,030220 oncology & carcinogenesis ,Cancer cell ,Colonic Neoplasms ,Cancer research ,Female ,fenretinide ,cancer stem cells ,cancer ,DNA Damage - Abstract
Fenretinide is a synthetic retinoid characterized by anticancer activity in preclinical models and favorable toxicological profile, but also by a low bioavailability that hindered its clinical efficacy in former clinical trials. We developed a new formulation of fenretinide complexed with 2-hydroxypropyl-beta-cyclodextrin (nanofenretinide) characterized by an increased bioavailability and therapeutic efficacy. Nanofenretinide was active in cell lines derived from multiple solid tumors, in primary spheroid cultures and in xenografts of lung and colorectal cancer, where it inhibited tumor growth independently from the mutational status of tumor cells. A global profiling of pathways activated by nanofenretinide was performed by reverse-phase proteomic arrays and lipid analysis, revealing widespread repression of the mTOR pathway, activation of apoptotic, autophagic and DNA damage signals and massive production of dihydroceramide, a bioactive lipid with pleiotropic effects on several biological processes. In cells that survived nanofenretinide treatment there was a decrease of factors involved in cell cycle progression and an increase in the levels of p16 and phosphorylated p38 MAPK with consequent block in G0 and early G1. The capacity of nanofenretinide to induce cancer cell death and quiescence, together with its elevated bioavailability and broad antitumor activity indicate its potential use in cancer treatment and chemoprevention.
- Published
- 2019
- Full Text
- View/download PDF
114. HDAC3 genetic and pharmacologic inhibition radiosensitizes fusion positive rhabdomyosarcoma by promoting DNA double-strand breaks.
- Author
-
Cassandri M, Porrazzo A, Pomella S, Noce B, Zwergel C, Aiello FA, Vulcano F, Milazzo L, Camero S, Pajalunga D, Spada M, Manzi V, Gravina GL, Codenotti S, Piccione M, Tomaciello M, Signore M, Barillari G, Marchese C, Fanzani A, De Angelis B, Quintarelli C, Vakoc CR, Chen EY, Megiorni F, Locatelli F, Valente S, Mai A, Rota R, and Marampon F
- Abstract
Radiotherapy (RT) plays a critical role in the management of rhabdomyosarcoma (RMS), the prevalent soft tissue sarcoma in childhood. The high risk PAX3-FOXO1 fusion-positive subtype (FP-RMS) is often resistant to RT. We have recently demonstrated that inhibition of class-I histone deacetylases (HDACs) radiosensitizes FP-RMS both in vitro and in vivo. However, HDAC inhibitors exhibited limited success on solid tumors in human clinical trials, at least in part due to the presence of off-target effects. Hence, identifying specific HDAC isoforms that can be targeted to radiosensitize FP-RMS is imperative. We, here, found that only HDAC3 silencing, among all class-I HDACs screened by siRNA, radiosensitizes FP-RMS cells by inhibiting colony formation. Thus, we dissected the effects of HDAC3 depletion using CRISPR/Cas9-dependent HDAC3 knock-out (KO) in FP-RMS cells, which resulted in Endoplasmatic Reticulum Stress activation, ERK inactivation, PARP1- and caspase-dependent apoptosis and reduced stemness when combined with irradiation compared to single treatments. HDAC3 loss-of-function increased DNA damage in irradiated cells augmenting H2AX phosphorylation and DNA double-strand breaks (DSBs) and counteracting irradiation-dependent activation of ATM and DNA-Pkcs as well as Rad51 protein induction. Moreover, HDAC3 depletion hampers FP-RMS tumor growth in vivo and maximally inhibits the growth of irradiated tumors compared to single approaches. We, then, developed a new HDAC3 inhibitor, MC4448, which showed specific cell anti-tumor effects and mirrors the radiosensitizing effects of HDAC3 depletion in vitro synergizing with ERKs inhibition. Overall, our findings dissect the pro-survival role of HDAC3 in FP-RMS and suggest HDAC3 genetic or pharmacologic inhibition as a new promising strategy to overcome radioresistance in this tumor., (© 2024. The Author(s).)
- Published
- 2024
- Full Text
- View/download PDF
115. Chlorpromazine overcomes temozolomide resistance in glioblastoma by inhibiting Cx43 and essential DNA repair pathways.
- Author
-
Matarrese P, Signore M, Ascione B, Fanelli G, Paggi MG, and Abbruzzese C
- Subjects
- Humans, Cell Line, Tumor, Gene Expression Regulation, Neoplastic drug effects, Drug Synergism, Brain Neoplasms drug therapy, Brain Neoplasms pathology, Brain Neoplasms genetics, Chlorpromazine pharmacology, Chlorpromazine therapeutic use, Temozolomide pharmacology, Temozolomide therapeutic use, Glioblastoma drug therapy, Glioblastoma pathology, Glioblastoma genetics, DNA Repair drug effects, Connexin 43 metabolism, Connexin 43 genetics, Drug Resistance, Neoplasm drug effects, Drug Resistance, Neoplasm genetics
- Abstract
Background: In the fight against GBM, drug repurposing emerges as a viable and time-saving approach to explore new treatment options. Chlorpromazine, an old antipsychotic medication, has recently arisen as a promising candidate for repositioning in GBM therapy in addition to temozolomide, the first-line standard of care. We previously demonstrated the antitumor efficacy of chlorpromazine and its synergistic effects with temozolomide in suppressing GBM cell malignant features in vitro. This prompted us to accomplish a Phase II clinical trial to evaluate the efficacy and safety of adding chlorpromazine to temozolomide in GBM patients with unmethylated MGMT gene promoter. In this in vitro study, we investigate the potential role of chlorpromazine in overcoming temozolomide resistance., Methods: In our experimental set, we analyzed Connexin-43 expression at both the transcriptional and protein levels in control- and chlorpromazine-treated GBM cells. DNA damage and subsequent repair were assessed by immunofluorescence of γ-H2AX and Reverse-Phase Protein microArrays in chlorpromazine treated GBM cell lines. To elucidate the relationship between DNA repair systems and chemoresistance, we analyzed a signature of DNA repair genes in GBM cells after treatment with chlorpromazine, temozolomide and Connexin-43 downregulation., Results: Chlorpromazine treatment significantly downregulated connexin-43 expression in GBM cells, consequently compromising connexin-dependent cellular resilience, and ultimately contributing to cell death. In line with this, we observed concordant post-translational modifications of molecular determinants involved in DNA damage and repair pathways. Our evaluation of DNA repair genes revealed that temozolomide elicited an increase, while chlorpromazine, as well as connexin-43 silencing, a decrease in DNA repair gene expression in GBM cells., Conclusions: Chlorpromazine potentiates the cytotoxic effects of the alkylating agent temozolomide through a mechanism involving downregulation of Cx43 expression and disruption of the cell cycle arrest essential for DNA repair processes. This finding suggests that chlorpromazine may be a potential therapeutic strategy to overcome TMZ resistance in GBM cells by inhibiting their DNA repair mechanisms., (© 2024. The Author(s).)
- Published
- 2024
- Full Text
- View/download PDF
116. Chlorpromazine affects glioblastoma bioenergetics by interfering with pyruvate kinase M2.
- Author
-
Abbruzzese C, Matteoni S, Matarrese P, Signore M, Ascione B, Iessi E, Gurtner A, Sacconi A, Ricci-Vitiani L, Pallini R, Pace A, Villani V, Polo A, Costantini S, Budillon A, Ciliberto G, and Paggi MG
- Subjects
- Humans, Chlorpromazine pharmacology, Chlorpromazine therapeutic use, Pyruvate Kinase metabolism, Cell Line, Tumor, Energy Metabolism, Glioblastoma pathology
- Abstract
Glioblastoma (GBM) is the most frequent and lethal brain tumor, whose therapeutic outcome - only partially effective with current schemes - places this disease among the unmet medical needs, and effective therapeutic approaches are urgently required. In our attempts to identify repositionable drugs in glioblastoma therapy, we identified the neuroleptic drug chlorpromazine (CPZ) as a very promising compound. Here we aimed to further unveil the mode of action of this drug. We performed a supervised recognition of the signal transduction pathways potentially influenced by CPZ via Reverse-Phase Protein microArrays (RPPA) and carried out an Activity-Based Protein Profiling (ABPP) followed by Mass Spectrometry (MS) analysis to possibly identify cellular factors targeted by the drug. Indeed, the glycolytic enzyme PKM2 was identified as one of the major targets of CPZ. Furthermore, using the Seahorse platform, we analyzed the bioenergetics changes induced by the drug. Consistent with the ability of CPZ to target PKM2, we detected relevant changes in GBM energy metabolism, possibly attributable to the drug's ability to inhibit the oncogenic properties of PKM2. RPE-1 non-cancer neuroepithelial cells appeared less responsive to the drug. PKM2 silencing reduced the effects of CPZ. 3D modeling showed that CPZ interacts with PKM2 tetramer in the same region involved in binding other known activators. The effect of CPZ can be epitomized as an inhibition of the Warburg effect and thus malignancy in GBM cells, while sparing RPE-1 cells. These preclinical data enforce the rationale that allowed us to investigate the role of CPZ in GBM treatment in a recent multicenter Phase II clinical trial., (© 2023. The Author(s).)
- Published
- 2023
- Full Text
- View/download PDF
117. Flow-dependent shear stress affects the biological properties of pericyte-like cells isolated from human dental pulp.
- Author
-
Bertani G, Di Tinco R, Bertoni L, Orlandi G, Pisciotta A, Rosa R, Rigamonti L, Signore M, Bertacchini J, Sena P, De Biasi S, Villa E, and Carnevale G
- Subjects
- Humans, Dental Pulp, Leukocytes, Mononuclear, Stem Cells, Pericytes, Endothelial Cells
- Abstract
Background: Human dental pulp stem cells represent a mesenchymal stem cell niche localized in the perivascular area of dental pulp and are characterized by low immunogenicity and immunomodulatory/anti-inflammatory properties. Pericytes, mural cells surrounding the endothelium of small vessels, regulate numerous functions including vessel growth, stabilization and permeability. It is well established that pericytes have a tight cross talk with endothelial cells in neoangiogenesis and vessel stabilization, which are regulated by different factors, i.e., microenvironment and flow-dependent shear stress. The aim of this study was to evaluate the effects of a pulsatile unidirectional flow in the presence or not of an inflammatory microenvironment on the biological properties of pericyte-like cells isolated from human dental pulp (hDPSCs)., Methods: Human DPSCs were cultured under both static and dynamic conditions with or without pre-activated peripheral blood mononuclear cells (PBMCs). Pulsatile unidirectional flow shear stress was generated by using a specific peristaltic pump. The angiogenic potential and inflammatory properties of hDPSCs were evaluated through reverse phase protein microarrays (RPPA), confocal immunofluorescence and western blot analyses., Results: Our data showed that hDPSCs expressed the typical endothelial markers, which were up-regulated after endothelial induction, and were able to form tube-like structures. RPPA analyses revealed that these properties were modulated when a pulsatile unidirectional flow shear stress was applied to hDPSCs. Stem cells also revealed a downregulation of the immune-modulatory molecule PD-L1, in parallel with an up-regulation of the pro-inflammatory molecule NF-kB. Immune-modulatory properties of hDPSCs were also reduced after culture under flow-dependent shear stress and exposure to an inflammatory microenvironment. This evidence was strengthened by the detection of up-regulated levels of expression of pro-inflammatory cytokines in PBMCs., Conclusions: In conclusion, the application of a pulsatile unidirectional flow shear stress induced a modulation of immunomodulatory/inflammatory properties of dental pulp pericyte-like cells., (© 2023. The Author(s).)
- Published
- 2023
- Full Text
- View/download PDF
118. Type I IFNs promote cancer cell stemness by triggering the epigenetic regulator KDM1B.
- Author
-
Musella M, Guarracino A, Manduca N, Galassi C, Ruggiero E, Potenza A, Maccafeo E, Manic G, Mattiello L, Soliman Abdel Rehim S, Signore M, Pietrosanto M, Helmer-Citterich M, Pallocca M, Fanciulli M, Bruno T, De Nicola F, Corleone G, Di Benedetto A, Ercolani C, Pescarmona E, Pizzuti L, Guidi F, Sperati F, Vitale S, Macchia D, Spada M, Schiavoni G, Mattei F, De Ninno A, Businaro L, Lucarini V, Bracci L, Aricò E, Ziccheddu G, Facchiano F, Rossi S, Sanchez M, Boe A, Biffoni M, De Maria R, Vitale I, and Sistigu A
- Subjects
- Anthracyclines metabolism, Anthracyclines therapeutic use, Female, Humans, Neoplastic Stem Cells metabolism, Neoplastic Stem Cells pathology, Breast Neoplasms drug therapy, Breast Neoplasms genetics, Breast Neoplasms pathology, Epigenesis, Genetic, Histone Demethylases metabolism, Interferon Type I metabolism
- Abstract
Cancer stem cells (CSCs) are a subpopulation of cancer cells endowed with high tumorigenic, chemoresistant and metastatic potential. Nongenetic mechanisms of acquired resistance are increasingly being discovered, but molecular insights into the evolutionary process of CSCs are limited. Here, we show that type I interferons (IFNs-I) function as molecular hubs of resistance during immunogenic chemotherapy, triggering the epigenetic regulator demethylase 1B (KDM1B) to promote an adaptive, yet reversible, transcriptional rewiring of cancer cells towards stemness and immune escape. Accordingly, KDM1B inhibition prevents the appearance of IFN-I-induced CSCs, both in vitro and in vivo. Notably, IFN-I-induced CSCs are heterogeneous in terms of multidrug resistance, plasticity, invasiveness and immunogenicity. Moreover, in breast cancer (BC) patients receiving anthracycline-based chemotherapy, KDM1B positively correlated with CSC signatures. Our study identifies an IFN-I → KDM1B axis as a potent engine of cancer cell reprogramming, supporting KDM1B targeting as an attractive adjunctive to immunogenic drugs to prevent CSC expansion and increase the long-term benefit of therapy., (© 2022. The Author(s).)
- Published
- 2022
- Full Text
- View/download PDF
119. An organoid model of colorectal circulating tumor cells with stem cell features, hybrid EMT state and distinctive therapy response profile.
- Author
-
De Angelis ML, Francescangeli F, Nicolazzo C, Signore M, Giuliani A, Colace L, Boe A, Magri V, Baiocchi M, Ciardi A, Scarola F, Spada M, La Torre F, Gazzaniga P, Biffoni M, De Maria R, and Zeuner A
- Subjects
- Biomarkers, Tumor metabolism, Epithelial-Mesenchymal Transition physiology, Humans, Organoids metabolism, Stem Cells metabolism, Colorectal Neoplasms drug therapy, Colorectal Neoplasms genetics, Neoplastic Cells, Circulating metabolism
- Abstract
Background: Circulating tumor cells (CTCs) are responsible for the metastatic dissemination of colorectal cancer (CRC) to the liver, lungs and lymph nodes. CTCs rarity and heterogeneity strongly limit the elucidation of their biological features, as well as preclinical drug sensitivity studies aimed at metastasis prevention., Methods: We generated organoids from CTCs isolated from an orthotopic CRC xenograft model. CTCs-derived organoids (CTCDOs) were characterized through proteome profiling, immunohistochemistry, immunofluorescence, flow cytometry, tumor-forming capacity and drug screening assays. The expression of intra- and extracellular markers found in CTCDOs was validated on CTCs isolated from the peripheral blood of CRC patients., Results: CTCDOs exhibited a hybrid epithelial-mesenchymal transition (EMT) state and an increased expression of stemness-associated markers including the two homeobox transcription factors Goosecoid and Pancreatic Duodenal Homeobox Gene-1 (PDX1), which were also detected in CTCs from CRC patients. Functionally, CTCDOs showed a higher migratory/invasive ability and a different response to pathway-targeted drugs as compared to xenograft-derived organoids (XDOs). Specifically, CTCDOs were more sensitive than XDOs to drugs affecting the Survivin pathway, which decreased the levels of Survivin and X-Linked Inhibitor of Apoptosis Protein (XIAP) inducing CTCDOs death., Conclusions: These results indicate that CTCDOs recapitulate several features of colorectal CTCs and may be used to investigate the features of metastatic CRC cells, to identify new prognostic biomarkers and to devise new potential strategies for metastasis prevention., (© 2022. The Author(s).)
- Published
- 2022
- Full Text
- View/download PDF
120. Reverse Phase Protein Arrays in cancer stem cells.
- Author
-
Signore M and Manganelli V
- Subjects
- Biomarkers, Neoplastic Stem Cells, Proteomics methods, Neoplasms metabolism, Protein Array Analysis methods
- Abstract
The scenario of proteogenomics is rapidly evolving and novel technologies are enabling comprehensive molecular exploration down to single cells. Likewise, digital (immuno-)assays are revolutionizing the field of biomarker detection and have reached the grade for population-level screenings with single-molecule sensitivity. Nonetheless, cost- and time-effective, high-throughput targeted phospho-proteomics at a preclinical stage still relies on ad hoc microarray platforms, such as the Reverse-Phase Protein microArrays (RPPA). Although this technique requires specific knowledge and equipment and different laboratories worldwide have implemented alternative methodological strategies, the application of RPPA to biomarker discovery has proven successful on diverse types of samples, including tissues and biological fluids as well as nanovesicles and in vitro cultured lines. Among these, cancer stem(-like) cells (CSC) represent an ideal experimental model system for preclinical discovery and definition of novel drug targets. The present methodological article provides the basic knowledge and steps on how to deploy an RPPA analysis with specific reference to an ideal experimental setup of drug testing on CSC., Competing Interests: Disclosures Authors have no conflicts of interest to disclose., (Copyright © 2022 Elsevier Inc. All rights reserved.)
- Published
- 2022
- Full Text
- View/download PDF
121. Elesclomol-induced increase of mitochondrial reactive oxygen species impairs glioblastoma stem-like cell survival and tumor growth.
- Author
-
Buccarelli M, D'Alessandris QG, Matarrese P, Mollinari C, Signore M, Cappannini A, Martini M, D'Aliberti P, De Luca G, Pedini F, Boe A, Biffoni M, Pallini R, and Ricci-Vitiani L
- Subjects
- Animals, Cell Line, Tumor, Cell Proliferation, Humans, Hydrazines pharmacology, Male, Mice, Mice, Inbred NOD, Oxidative Stress, Reactive Oxygen Species, Brain Neoplasms drug therapy, Cell Survival drug effects, Glioblastoma drug therapy, Hydrazines therapeutic use
- Abstract
Background: Glioblastoma (GBM) is the most common and aggressive primary malignant brain tumor in adults, characterized by a poor prognosis mainly due to recurrence and therapeutic resistance. It has been widely demonstrated that glioblastoma stem-like cells (GSCs), a subpopulation of tumor cells endowed with stem-like properties is responsible for tumor maintenance and progression. Moreover, it has been demonstrated that GSCs contribute to GBM-associated neovascularization processes, through different mechanisms including the transdifferentiation into GSC-derived endothelial cells (GdECs)., Methods: In order to identify druggable cancer-related pathways in GBM, we assessed the effect of a selection of 349 compounds on both GSCs and GdECs and we selected elesclomol (STA-4783) as the most effective agent in inducing cell death on both GSC and GdEC lines tested., Results: Elesclomol has been already described to be a potent oxidative stress inducer. In depth investigation of the molecular mechanisms underlying GSC and GdEC response to elesclomol, confirmed that this compound induces a strong increase in mitochondrial reactive oxygen species (ROS) in both GSCs and GdECs ultimately leading to a non-apoptotic copper-dependent cell death. Moreover, combined in vitro treatment with elesclomol and the alkylating agent temozolomide (TMZ) enhanced the cytotoxicity compared to TMZ alone. Finally, we used our experimental model of mouse brain xenografts to test the combination of elesclomol and TMZ and confirmed their efficacy in vivo., Conclusions: Our results support further evaluation of therapeutics targeting oxidative stress such as elesclomol with the aim of satisfying the high unmet medical need in the management of GBM., (© 2021. The Author(s).)
- Published
- 2021
- Full Text
- View/download PDF
122. Control of replication stress and mitosis in colorectal cancer stem cells through the interplay of PARP1, MRE11 and RAD51.
- Author
-
Manic G, Musella M, Corradi F, Sistigu A, Vitale S, Soliman Abdel Rehim S, Mattiello L, Malacaria E, Galassi C, Signore M, Pallocca M, Scalera S, Goeman F, De Nicola F, Guarracino A, Pennisi R, Antonangeli F, Sperati F, Baiocchi M, Biffoni M, Fanciulli M, Maugeri-Saccà M, Franchitto A, Pichierri P, De Maria R, and Vitale I
- Subjects
- Antineoplastic Agents pharmacology, Cell Line, Tumor, Colorectal Neoplasms genetics, DNA Replication drug effects, Humans, MRE11 Homologue Protein genetics, Neoplastic Stem Cells metabolism, Poly (ADP-Ribose) Polymerase-1 genetics, Rad51 Recombinase genetics, Colorectal Neoplasms drug therapy, MRE11 Homologue Protein drug effects, Mitosis drug effects, Neoplastic Stem Cells drug effects, Poly (ADP-Ribose) Polymerase-1 drug effects, Rad51 Recombinase drug effects
- Abstract
Cancer stem cells (CSCs) are tumor subpopulations driving disease development, progression, relapse and therapy resistance, and their targeting ensures tumor eradication. CSCs display heterogeneous replication stress (RS), but the functionality/relevance of the RS response (RSR) centered on the ATR-CHK1 axis is debated. Here, we show that the RSR is efficient in primary CSCs from colorectal cancer (CRC-SCs), and describe unique roles for PARP1 and MRE11/RAD51. First, we demonstrated that PARP1 is upregulated in CRC-SCs resistant to several replication poisons and RSR inhibitors (RSRi). In these cells, PARP1 modulates replication fork speed resulting in low constitutive RS. Second, we showed that MRE11 and RAD51 cooperate in the genoprotection and mitosis execution of PARP1-upregulated CRC-SCs. These roles represent therapeutic vulnerabilities for CSCs. Indeed, PARP1i sensitized CRC-SCs to ATRi/CHK1i, inducing replication catastrophe, and prevented the development of resistance to CHK1i. Also, MRE11i + RAD51i selectively killed PARP1-upregulated CRC-SCs via mitotic catastrophe. These results provide the rationale for biomarker-driven clinical trials in CRC using distinct RSRi combinations.
- Published
- 2021
- Full Text
- View/download PDF
123. Diagnostic and prognostic potential of the proteomic profiling of serum-derived extracellular vesicles in prostate cancer.
- Author
-
Signore M, Alfonsi R, Federici G, Nanni S, Addario A, Bertuccini L, Aiello A, Di Pace AL, Sperduti I, Muto G, Giacobbe A, Collura D, Brunetto L, Simone G, Costantini M, Crinò L, Rossi S, Tabolacci C, Diociaiuti M, Merlino T, Gallucci M, Sentinelli S, Papalia R, De Maria R, and Bonci D
- Subjects
- Adult, Aged, Cell Line, Tumor, Extracellular Vesicles ultrastructure, Humans, Male, Middle Aged, Predictive Value of Tests, Prostatic Neoplasms ultrastructure, Protein Array Analysis, Reproducibility of Results, Retrospective Studies, Biomarkers, Tumor blood, Extracellular Vesicles metabolism, Neoplasm Proteins blood, Prostatic Neoplasms blood, Proteome, Proteomics
- Abstract
Extracellular vesicles (EVs) and their cargo represent an intriguing source of cancer biomarkers for developing robust and sensitive molecular tests by liquid biopsy. Prostate cancer (PCa) is still one of the most frequent and deadly tumor in men and analysis of EVs from biological fluids of PCa patients has proven the feasibility and the unprecedented potential of such an approach. Here, we exploited an antibody-based proteomic technology, i.e. the Reverse-Phase Protein microArrays (RPPA), to measure key antigens and activated signaling in EVs isolated from sera of PCa patients. Notably, we found tumor-specific protein profiles associated with clinical settings as well as candidate markers for EV-based tumor diagnosis. Among others, PD-L1, ERG, Integrin-β5, Survivin, TGF-β, phosphorylated-TSC2 as well as partners of the MAP-kinase and mTOR pathways emerged as differentially expressed endpoints in tumor-derived EVs. In addition, the retrospective analysis of EVs from a 15-year follow-up cohort generated a protein signature with prognostic significance. Our results confirm that serum-derived EV cargo may be exploited to improve the current diagnostic procedures while providing potential prognostic and predictive information. The approach proposed here has been already applied to tumor entities other than PCa, thus proving its value in translational medicine and paving the way to innovative, clinically meaningful tools.
- Published
- 2021
- Full Text
- View/download PDF
124. The clinical value of patient-derived glioblastoma tumorspheres in predicting treatment response.
- Author
-
D'Alessandris QG, Biffoni M, Martini M, Runci D, Buccarelli M, Cenci T, Signore M, Stancato L, Olivi A, De Maria R, Larocca LM, Ricci-Vitiani L, and Pallini R
- Subjects
- Aged, Antineoplastic Agents, Alkylating therapeutic use, Brain Neoplasms pathology, Chemoradiotherapy methods, Dacarbazine analogs & derivatives, Dacarbazine therapeutic use, Disease-Free Survival, Female, Glioblastoma mortality, Glioblastoma pathology, Humans, Male, Middle Aged, Temozolomide, Brain Neoplasms therapy, Glioblastoma therapy
- Abstract
Background: Advances from glioma stemlike cell (GSC) research, though increasing our knowledge of glioblastoma (GBM) biology, do not influence clinical decisions yet. We explored the translational power of GSC-enriched cultures from patient-derived tumorspheres (TS) in predicting treatment response., Methods: The relationship between TS growth and clinical outcome was investigated in 52 GBMs treated with surgical resection followed by radiotherapy and temozolomide (TMZ). The effect on TS of radiation (6 to 60 Gy) and of TMZ (3.9 μM to 1 mM) was related with patients' survival., Results: Generation of TS was an independent factor for poor overall survival (OS) and poor progression-free survival (PFS) (P < .0001 and P = .0010, respectively). Growth rate and clonogenicity of TS predicted poor OS. In general, TS were highly resistant to both radiation and TMZ. Resistance to TMZ was stronger in TS with high clonogenicity and fast growth (P < .02). Shorter PFS was associated with radiation LD50 (lethal dose required to kill 50% of TS cells) >12 Gy of matched TS (P = .0484). A direct relationship was found between sensitivity of TS to TMZ and patients' survival (P = .0167 and P = .0436 for OS and PFS, respectively). Importantly, values for TMZ half-maximal inhibitory concentration <50 μM, which are in the range of plasma levels achieved in vivo, identified cases with longer OS and PFS (P = .0020 and P = .0016, respectively)., Conclusions: Analysis of TS holds translational relevance by predicting the response of parent tumors to radiation and, particularly, to TMZ. Dissecting the clonogenic population from proliferating progeny in TS can guide therapeutic strategies to a more effective drug selection and treatment duration., (© The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com)
- Published
- 2017
- Full Text
- View/download PDF
125. Antibody Validation by Western Blotting.
- Author
-
Signore M, Manganelli V, and Hodge A
- Subjects
- Animals, Humans, Antibody Specificity, Blotting, Western methods
- Abstract
Validation of antibodies is an integral part of translational research, particularly for biomarker discovery. Assaying the specificity of the reagent (antibody) and confirming the identity of the protein biomarker is of critical importance prior to implementing any biomarker in clinical studies, and the lack of such quality control tests may result in unexpected and/or misleading results.Antibody validation is the procedure in which a single antibody is thoroughly assayed for sensitivity and specificity. Although a plethora of commercial antibodies exist, antibody specificity must be extensively demonstrated using diverse complex biological samples, rather than purified recombinant proteins, prior to use in clinical translational research. In the simplest iteration, antibody specificity is determined by the presence of a single band in a complex biological sample, at the expected molecular weight, on a Western blot.To date, numerous Western blotting procedures are available, based on either manual or automated systems and spanning the spectrum of single blots to multiplex blots. X-ray film is still employed in many research laboratories, but digital imaging has become a gold standard in immunoblotting. The basic principles of Western blotting are (a) separation of protein mixtures by gel electrophoresis, (b) transfer of the proteins to a blot, (c) probing the blot for a protein or proteins of interest, and (d) subsequent detection of the protein by chemiluminescent, fluorescent, or colorimetric methods. This chapter focuses on the chemiluminescent detection of proteins using a manual Western blotting system and a vacuum-enhanced detection system (SNAP i.d.™, Millipore).
- Published
- 2017
- Full Text
- View/download PDF
126. Antibody validation by Western blotting.
- Author
-
Signore M and Reeder KA
- Subjects
- Animals, Antibodies metabolism, Antibody Specificity immunology, Humans, Proteins analysis, Proteins immunology, Reproducibility of Results, Sensitivity and Specificity, Antibodies immunology, Blotting, Western methods
- Abstract
Validation of antibodies is an integral part of translational research, particularly for biomarker discovery. Validation is essential to show the specificity of the reagent (antibody) and to confirm the identity of the protein biomarker, prior to implementing the biomarker in clinical studies.Antibody validation is the procedure in which a single antibody is thoroughly assayed for sensitivity and specificity. Although a plethora of commercial antibodies exist, antibody specificity must be thoroughly demonstrated using a complex biological sample, rather than a recombinant protein, prior to use in clinical translational research. In the simplest iteration, antibody specificity is determined by the presence of a single band in a complex biological sample, at the expected molecular weight, on a western blot.Numerous western blotting procedures are available, spanning the spectrum of single blots to multiplex blots, with images and quantitation generated by manual or automated systems. The basic principles of western blotting are (a) separation of protein mixtures by gel electrophoresis, (b) transfer of the proteins to a blot, (c) probing the blot for a protein or proteins of interest, and (d) subsequent detection of the protein by chemiluminescent, fluorescent, or colorimetric methods. This chapter focuses on the chemiluminescent detection of proteins using a manual western blotting system and a vacuum-enhanced detection system (SNAP i.d.™, Millipore).
- Published
- 2012
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.