Publications by authors named "Clara Nervi"

51 Publications

Molecular imaging in immuno-oncology: current status and translational perspectives.

Expert Rev Mol Diagn 2020 12 7;20(12):1199-1211. Epub 2020 Dec 7.

Department of Nuclear Medicine, Santa Maria Goretti Hospital, AUSL , Latina, Italy.

: Only 20-40% of patients respond to therapy with immune checkpoint inhibitors (ICIs). Therefore, the early identification of subjects that can benefit from such therapeutic regimen is mandatory. : The immunobiological mechanisms of ICIs are briefly illustrated. Furthermore, the limitations of traditional radiological approaches are covered. Then, the pros and cons of molecular imaging through positron emission computed tomography (PET/CT) are reviewed, with a particular focus on f-fluorodeoxyglucose (F-FDG) and PET-derived metabolic parameters. Lastly, translational perspective of radiopharmaceuticals others than F-FDG such as zirconium (Zr) or fluorine-18 (F) labeled monoclonal antibodies (e.g.Zr-atezolizumab, Zr-nivolumab) binding to specific biomarkers are discussed. : Molecular imaging presents a prominent role for the management of oncological patients treated with ICIs. Preliminary clinical data indicate that PET/CT with F-FDG is useful for assessing the response to treatment and for the imaging of immune-related adverse effects. Nevertheless, the methodological approach (iPERCIST, PERCIMT, or others) to be used for an optimal diagnostic accuracy and patients' evaluation is still a debated issue. PET/CT with radioligands directed toward ICIs biomarkers, although is still in a translational phase, holds the promise of accurately predicting the response to treatment and revealing the acquired resistance to immunotherapy.
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http://dx.doi.org/10.1080/14737159.2020.1854090DOI Listing
December 2020

Transcriptional and Metabolic Dissection of ATRA-Induced Granulocytic Differentiation in NB4 Acute Promyelocytic Leukemia Cells.

Cells 2020 11 5;9(11). Epub 2020 Nov 5.

Dipartimento di Scienze, Sezione di Scienze e Tecnologie Biomediche, Università Roma Tre, 00146 Roma, Italy.

Acute promyelocytic leukemia (APL) is a hematological disease characterized by a balanced reciprocal translocation that leads to the synthesis of the oncogenic fusion protein PML-RARα. APL is mainly managed by a differentiation therapy based on the administration of all- retinoic acid (ATRA) and arsenic trioxide (ATO). However, therapy resistance, differentiation syndrome, and relapses require the development of new low-toxicity therapies based on the induction of blasts differentiation. In keeping with this, we reasoned that a better understanding of the molecular mechanisms pivotal for ATRA-driven differentiation could definitely bolster the identification of new therapeutic strategies in APL patients. We thus performed an in-depth high-throughput transcriptional profile analysis and metabolic characterization of a well-established APL experimental model based on NB4 cells that represent an unevaluable tool to dissect the complex mechanism associated with ATRA-induced granulocytic differentiation. Pathway-reconstruction analysis using genome-wide transcriptional data has allowed us to identify the activation/inhibition of several cancer signaling pathways (e.g., inflammation, immune cell response, DNA repair, and cell proliferation) and master regulators (e.g., transcription factors, epigenetic regulators, and ligand-dependent nuclear receptors). Furthermore, we provide evidence of the regulation of a considerable set of metabolic genes involved in cancer metabolic reprogramming. Consistently, we found that ATRA treatment of NB4 cells drives the activation of aerobic glycolysis pathway and the reduction of OXPHOS-dependent ATP production. Overall, this study represents an important resource in understanding the molecular "portfolio" pivotal for APL differentiation, which can be explored for developing new therapeutic strategies.
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http://dx.doi.org/10.3390/cells9112423DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7716236PMC
November 2020

Aptamer-based technology for radionuclide targeted imaging and therapy: a promising weapon against cancer.

Expert Rev Med Devices 2020 Aug 27;17(8):751-758. Epub 2020 Jul 27.

Department of Medical and Surgical Sciences and Biotechnology, University of Rome "La Sapienza" , Latina, Italy.

Introduction: aptamers are short artificial, single-strand oligonucleotide sequences (DNA, RNA or modified RNA), capable of binding to biological molecules with high affinity and specificity. Due to their relatively low cost of production and scarce immunogenicity, many efforts have been made to produce aptamers directed against specific molecular targets, such as receptors or transporters overexpressed by malignancies.

Areas Covered: the technological approaches for generating aptamers are reviewed. Furthermore, the applications of radiolabeled aptamers for the imaging of several oncological biomarkers through single photon emission computed tomography (SPECT) or positron emission tomography (PET), are covered. Lastly, targeted therapy based on the utilization of aptamers labeled with radionuclides emitting beta particles is discussed, with particular emphasis to the oncological perspectives.

Expert Opinion: The main limitation of radiolabeled aptamers is represented by their sensitivity to endogenous nuclease, so that several strategies have been developed to increase the stability of these compounds. Although the applications of aptamers are still in a preliminary and pre-clinical phase, it is reasonable to hypothesize that this technology will play a major role for personalized medicine in the next years.
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http://dx.doi.org/10.1080/17434440.2020.1796633DOI Listing
August 2020

Ruxolitinib binding to human serum albumin: bioinformatics, biochemical and functional characterization in JAK2V617F cell models.

Sci Rep 2019 11 8;9(1):16379. Epub 2019 Nov 8.

Interdepartmental Laboratory for Electron Microscopy, Roma Tre University, I-00146, Roma, Italy.

Ruxolitinib is a type I JAK inhibitor approved by FDA for targeted therapy of Philadelphia-negative myeloproliferative neoplasms (MPNs), all characterized by mutations activating the JAK2/STAT signaling pathway. Treatment with ruxolitinib improves constitutional symptoms and splenomegaly. However, patients can become resistant to treatment and chronic therapy has only a mild effect on molecular/pathologic remissions. Drugs interaction with plasma proteins, i.e. human serum albumin (HSA), is an important factor affecting the intensity and duration of their pharmacological actions. Here, the ruxolitinib recognition by the fatty acid binding sites (FAs) 1, 6, 7, and 9 of HSA has been investigated from the bioinformatics, biochemical and/or biological viewpoints. Docking simulations indicate that ruxolitinib binds to multiple sites of HSA. Ruxolitinib binds to the FA1 and FA7 sites of HSA with high affinity (K = 3.1 μM and 4.6 μM, respectively, at pH 7.3 and 37.0 °C). Moreover, HSA selectively blocks, in a dose dependent manner, the cytotoxic activity of ruxolitinib in JAK2V617F cellular models for MPN, in vitro. Furthermore this event is accompanied by changes in the cell cycle, p27 and cyclin D3 levels, and JAK/STAT signaling. Given the high plasma concentration of HSA, ruxolitinib trapping may be relevant in vivo.
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http://dx.doi.org/10.1038/s41598-019-52852-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6841977PMC
November 2019

Are DNA damage response kinases a target for the differentiation treatment of acute myeloid leukemia?

IUBMB Life 2018 11 8;70(11):1057-1066. Epub 2018 Oct 8.

Department of Sciences, Roma Tre University, Roma, Italy.

Acute myeloid leukemia (AML) is a genetically heterogeneous malignancy characterized by the expansion of hematopoietic stem/progenitor cells (HPCs) blocked at different stages of maturation/differentiation. The poor outcome of AMLs necessitates therapeutic improvement. In AML, genes encoding for myeloid transcription factors, signaling receptors regulating cell proliferation, and epigenetic modifiers can be mutated by somatically acquired genetic mutations or altered by chromosomal translocations. These mutations modify chromatin organization at genes sites regulating HPCs proliferation, terminal differentiation, and DNA repair, contributing to the development and progression of the disease. The reversibility of the epigenetic modifications by drug treatment makes epigenetic changes attractive targets for AML therapeutic intervention. Recent findings underline increased DNA damage and abnormalities in the DNA damage response (DDR) as a critical feature of AML blasts. The DDR preserves cell integrity and must be tightly coordinated with DNA methylation and chromatin remodeling to ensure the accessibility to the DNA of transcription factors and repair enzymes. A crucial role in these events is played by the ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3-related protein (ATR) kinases, which are hyperactive in AML. Based on these findings, we hypothesize the inhibition of DNA damage kinases as an alternative or complementary strategy for the differentiation treatment of AML as it leads to a reduced ability to repair the DNA damage, and to the inhibition of specific epigenetic modifiers whose function is altered in leukemic cells. © 2018 IUBMB Life, 70(11):1057-1066, 2018.
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http://dx.doi.org/10.1002/iub.1918DOI Listing
November 2018

Eicosapentaenoic acid modulates the synergistic action of CREB1 and ID/E2A family members in the rat pup brain and mouse embryonic stem cells.

Biochim Biophys Acta Gene Regul Mech 2017 Aug 27;1860(8):870-884. Epub 2017 Jun 27.

Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Rome, Italy. Electronic address:

The aim of this study was to investigate the molecular mechanism by which eicosapentaenoic acid (EPA) may exert neuroprotective effects through an "EPA-cyclic AMP response element-binding protein (CREB)" signaling pathway. The current study reveals that EPA modulates the exquisite interplay of interaction of CREB1 with the inhibitor of DNA binding (ID) and E2A family members, thereby delivering mechanistic insights into specific neural differentiation program. In this scenario, our work provides evidence for the capability of CREB1 to sequester ID:E2A family members in brain tissues and neural differentiating mouse embryonic stem cells (mESCs) through formation of a [CREB1]:ID2:E47 tetrameric complex.In essence, the molecular function of CREB1 is to dynamically regulate the location-specific assembly or disassembly of basic-helix-loop-helix (bHLH):HLH protein complexes to mediate the activation of neural/glial target genes. Together, these findings support the one-to-many binding mechanism of CREB1 and indicate that EPA treatment potentiates the integration of CREB dependent signaling with HLH/bHLH transcriptional network, adding specificity to the CREB1-mediated gene regulation during neural/glial differentiation. Our current research on the EPA-CREB axis could reveal new molecular targets for treating neurogenerative disease.
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http://dx.doi.org/10.1016/j.bbagrm.2017.06.002DOI Listing
August 2017

A novel epigenetic mini-circuitry contributes to t(8;21) leukaemogenesis.

EMBO Mol Med 2017 07;9(7):933-949

Department of Haematology, Chinese PLA General Hospital, Beijing, China

DNA methylation patterns are frequently deregulated in t(8;21) acute myeloid leukaemia (AML), but little is known of the mechanisms by which specific gene sets become aberrantly methylated. Here, we found that the promoter DNA methylation signature of t(8;21) AML blasts differs from that of t(8;21) AMLs. This study demonstrated that a novel hypermethylated zinc finger-containing protein, THAP10, is a target gene and can be epigenetically suppressed by AML1-ETO at the transcriptional level in t(8;21) AML. Our findings also show that is a target of that can be epigenetically activated by the AML1-ETO recruiting co-activator p300. In this study, we demonstrated that epigenetic suppression of is the mechanistic link between AML1-ETO fusion proteins and tyrosine kinase cascades. In addition, we showed that THAP10 is a nuclear protein that inhibits myeloid proliferation and promotes differentiation both and Altogether, our results revealed an unexpected and important epigenetic mini-circuit of in t(8;21) AML, in which epigenetic suppression of predicts a poor clinical outcome and represents a novel therapeutic target.
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http://dx.doi.org/10.15252/emmm.201607180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5577530PMC
July 2017

A minicircuitry of microRNA-9-1 and RUNX1-RUNX1T1 contributes to leukemogenesis in t(8;21) acute myeloid leukemia.

Int J Cancer 2017 Feb 10;140(3):653-661. Epub 2016 Nov 10.

Department of Hematology, Chinese PLA General Hospital, Beijing, 100853, China.

MicroRNA-9-1(miR-9-1) plays an important role in the mechanism that regulates the lineage fate of differentiating hematopoietic cells. Recent studies have shown that miR-9-1 is downregulated in t (8; 21) AML. However, the pathogenic mechanisms underlying miR-9-1 downregulation and the RUNX1-RUNX1T1 fusion protein, generated from the translocation of t (8; 21) in AML, remain unclear. RUNX1-RUNX1T1 can induce leukemogenesis through resides in and functions as a stable RUNX1-RUNX1T1-containing transcription factor complex. In this study, we demonstrate that miR-9-1 expression increases significantly after the treatment of RUNX1-RUNX1T1 (+) AML cell lines with decitabine (a DNMT inhibitor) and trichostatin A (an HDAC inhibitor). In addition, we show that RUNX1-RUNX1T1 triggers the heterochromatic silencing of miR-9-1 by binding to RUNX1-binding sites in the promoter region of miR-9-1 and recruiting chromatin-remodeling enzymes, DNMTs, and HDACs, contributing to hypermethylation of miR-9-1 in t (8; 21) AML. Furthermore, because RUNX1, RUNX1T1, and RUNX1-RUNX1T1 are all regulated by miR-9-1, the silencing of miR-9-1 enhances the oncogenic activity of these genes. Besides, overexpression of miR-9-1 induces differentiation and inhibits proliferation in t (8; 21) AML cell lines. In conclusion, our results indicate a feedback circuitry involving miR-9-1 and RUNX1-RUNX1T1, contributing to leukemogenesis in RUNX1-RUNX1T1 (+) AML cell lines.
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http://dx.doi.org/10.1002/ijc.30481DOI Listing
February 2017

PML/RARa inhibits PTEN expression in hematopoietic cells by competing with PU.1 transcriptional activity.

Oncotarget 2016 Oct;7(41):66386-66397

Department of Biomedicine and Prevention, University of Rome "Tor Vergata", Rome, Italy.

Acute promyelocitic leukemia (APL) is characterized by the pathognomonic presence in leukemic blasts of the hybrid protein PML/RARA, that acts as a transcriptional repressor impairing the expression of genes that are critical to myeloid differentiation. Here, we show that primary blasts from APL patients express lower levels of the oncosuppressor protein PTEN, as compared to blast cells from other AML subtypes or normal bone marrow, and demonstrate that PML-RARA directly inhibits PTEN expression. We show that All-Trans Retinoic Acid (ATRA) triggers in APL cells an active chromatin status at the core regulatory region of the PTEN promoter, that allows the binding of the myeloid-regulating transcription factor PU.1, and, in turn, the transcriptional induction of PTEN. ATRA, via PML/RARA degradation, also promotes PTEN nuclear re-localization and decreases expression of the PTEN target Aurora A kinase. In conclusion, our findings support the notion that PTEN is one of the primary targets of PML/RARA in APL.
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http://dx.doi.org/10.18632/oncotarget.11964DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5341808PMC
October 2016

Epigenetic treatment of solid tumours: a review of clinical trials.

Clin Epigenetics 2015 10;7:127. Epub 2015 Dec 10.

Department of Medical and Surgical Sciences and Biotechnology, University of Rome "la Sapienza", Corso della Repubblica, 97, 04100 Latina, Italy.

Epigenetic treatment has been approved by regulatory agencies for haematological malignancies. The success observed in cutaneous lymphomas represents a proof of principle that similar results may be obtained in solid tumours. Several agents that interfere with DNA methylation-demethylation and histones acetylation/deacetylation have been studied, and some (such as azacytidine, decitabine, valproic acid and vorinostat) are already in clinical use. The aim of this review is to provide a brief overview of the molecular events underlying the antitumour effects of epigenetic treatments and to summarise data available on clinical trials that tested the use of epigenetic agents against solid tumours. We not only list results but also try to indicate how the proper evaluation of this treatment might result in a better selection of effective agents and in a more rapid development. We divided compounds in demethylating agents and HDAC inhibitors. For each class, we report the antitumour activity and the toxic side effects. When available, we describe plasma pharmacokinetics and pharmacodynamic evaluation in tumours and in surrogate tissues (generally white blood cells). Epigenetic treatment is a reality in haematological malignancies and deserves adequate attention in solid tumours. A careful consideration of available clinical data however is required for faster drug development and possibly to re-evaluate some molecules that were perhaps discarded too early.
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http://dx.doi.org/10.1186/s13148-015-0157-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4676165PMC
December 2015

Retinoic acid receptors: from molecular mechanisms to cancer therapy.

Mol Aspects Med 2015 Feb 25;41:1-115. Epub 2014 Dec 25.

Department of Medical and Surgical Sciences and Biotechnologies, University of Roma "La Sapienza", Corso della Repubblica 79, Latina I-04100. Electronic address:

Retinoic acid (RA), the major bioactive metabolite of retinol or vitamin A, induces a spectrum of pleiotropic effects in cell growth and differentiation that are relevant for embryonic development and adult physiology. The RA activity is mediated primarily by members of the retinoic acid receptor (RAR) subfamily, namely RARα, RARβ and RARγ, which belong to the nuclear receptor (NR) superfamily of transcription factors. RARs form heterodimers with members of the retinoid X receptor (RXR) subfamily and act as ligand-regulated transcription factors through binding specific RA response elements (RAREs) located in target genes promoters. RARs also have non-genomic effects and activate kinase signaling pathways, which fine-tune the transcription of the RA target genes. The disruption of RA signaling pathways is thought to underlie the etiology of a number of hematological and non-hematological malignancies, including leukemias, skin cancer, head/neck cancer, lung cancer, breast cancer, ovarian cancer, prostate cancer, renal cell carcinoma, pancreatic cancer, liver cancer, glioblastoma and neuroblastoma. Of note, RA and its derivatives (retinoids) are employed as potential chemotherapeutic or chemopreventive agents because of their differentiation, anti-proliferative, pro-apoptotic, and anti-oxidant effects. In humans, retinoids reverse premalignant epithelial lesions, induce the differentiation of myeloid normal and leukemic cells, and prevent lung, liver, and breast cancer. Here, we provide an overview of the biochemical and molecular mechanisms that regulate the RA and retinoid signaling pathways. Moreover, mechanisms through which deregulation of RA signaling pathways ultimately impact on cancer are examined. Finally, the therapeutic effects of retinoids are reported.
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http://dx.doi.org/10.1016/j.mam.2014.12.003DOI Listing
February 2015

RARs and microRNAs.

Subcell Biochem 2014 ;70:151-79

Department of Medical-Surgical Sciences and Biotechnologies, University "La Sapienza", Rome, Italy,

MicroRNA MicroRNA s (miRNAs) are small noncoding RNAs acting as endogenous regulators of gene expression. Their discovery is one of the major recent breakthroughs in molecular biology. miRNAs establish a multiplicity of relationships with target mRNAs and exert pleiotropic biological effects in many cell physiological pathways during development and adult life. The dynamic nature of gene expression regulation by Retinoic Acid Retinoic acid (RA) is consistent with an extensive functional interplay with miRNA activities. In fact, RA regulates the expression of many different miRNAs, thus suggesting a relevant function of miRNAs in RA-controlled gene expression programmes. miRNAs have been extensively studied as targets and mediators of the biological activity of RA during embryonic development as well as in normal and neoplastic cells. However, relatively few studies have experimentally explored the direct contribution of miRNA function to the RA signalling pathway. Here, we provide an overview of the mechanistic aspects that allow miRNA biogenesis, functional activation and regulation, focusing on recent evidence that highlights a functional interplay between miRNAs and RA-regulated molecular networks. We report examples of tissue-specific roles of miRNAs modulated by RA in stem cell pluripotency maintenance and regeneration, embryonic development, hematopoietic and neural differentiation, and other biological model systems, underlining their role in disease pathogenesis. We also address novel areas of research linking the RA signalling pathway to the nuclear activity of miRNAs.
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http://dx.doi.org/10.1007/978-94-017-9050-5_8DOI Listing
November 2014

AML1-ETO triggers epigenetic activation of early growth response gene l, inducing apoptosis in t(8;21) acute myeloid leukemia.

FEBS J 2014 Feb 10;281(4):1123-31. Epub 2014 Jan 10.

Department of Hematology, Chinese PLA General Hospital, Beijing, China; Nankai University School of Medicine, Tianjin, China.

The t(8;21)(q22;q22) translocation is the most common chromosomal translocation in acute myeloid leukemia (AML), and it gives rise to acute myeloid gene 1 (AML1)-myeloid transforming gene 8 (ETO)-positive AML, which has a relatively favorable prognosis. However, the molecular mechanism related to a favorable prognosis in AML1-ETO-positive AML is still not fully understood. Our results show that the AML1-ETO fusion protein triggered activation of early growth response gene l (EGR1) by binding at AML1-binding sites on the EGR1 promoter and, subsequently, recruiting acetyltransferase P300, which is known to acetylate histones. However, AML1-ETO could not recruit DNA methyltransferases and histone deacetylases; therefore, EGR1 expression was affected by histone acetylation but not by DNA methylation. Both transcription and translation of EGR1 were higher in AML1-ETO-positive AML cell lines than in AML1-ETO-negative AML cell lines, owing to acetylation. Furthermore, when AML1-ETO-positive AML cell lines were treated with C646 (P300 inhibitor) and trichostatin A (histone deacetylase inhibitor), EGR1 expression was significantly decreased and increased, respectively. In addition, treatment with 5-azacytidine (methyltransferase inhibitor) did not cause any significant change in EGR1 expression. Overexpression of EGR1 inhibited cell proliferation and promoted apoptosis, and EGR1 knockout promoted cell proliferation. Thus, EGR1 could be a novel prognostic factor for a favorable outcome in AML1-ETO-positive AML. The results of our study may explain the molecular mechanisms underlying the favorable prognosis in AML1-ETO-positive AML.
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http://dx.doi.org/10.1111/febs.12673DOI Listing
February 2014

A highly specific q-RT-PCR assay to address the relevance of the JAK2WT and JAK2V617F expression levels and control genes in Ph-negative myeloproliferative neoplasms.

Ann Hematol 2014 Apr 31;93(4):609-16. Epub 2013 Oct 31.

Department of Medico-Surgical Sciences and Biotechnologies, University of Rome "Sapienza" Polo Pontino, , Latina, Italy.

In Ph- myeloproliferative neoplasms, the quantification of the JAK2V617F transcripts may provide some advantages over the DNA allele burden determination. We developed a q-RT-PCR to assess the JAK2WT and JAK2V617F mRNA expression in 105 cases (23 donors, 13 secondary polycythemia, 22 polycythemia vera (PV), 38 essential thrombocythemia (ET), and 9 primary myelofibrosis (PMF)). Compared with the standard allele-specific oligonucleotide (ASO)-PCR technique, our assay showed a 100 % concordance rate detecting the JAK2V617F mutation in 22/22 PV (100 %), 29/38 (76.3 %) ET, and 5/9 (55.5 %) PMF cases, respectively. The sensitivity of the assay was 0.01 %. Comparing DNA and RNA samples, we found that the JAK2V617F mutational ratios were significantly higher at the RNA level both in PV (p = 0.005) and ET (p = 0.001) samples. In PV patients, JAK2WT expression levels positively correlated with the platelets (PLTs) (p = 0.003) whereas a trend to negative correlation was observed with the Hb levels (p = 0.051). JAK2V617F-positive cases showed the lowest JAK2WT and ABL1 mRNA expression levels. In all the samples, the expression pattern of beta-glucoronidase (GUSB) was more homogeneous than that of ABL1 or β2 microglobulin (B2M). Using GUSB as normalizator gene, a significant increase of the JAK2V617F mRNA levels was seen in two ET patients at time of progression to PV. In conclusion, the proposed q-RT-PCR is a sensitive and accurate method to quantify the JAK2 mutational status that can also show clinical correlations suggesting the impact of the residual amount of the JAK2WT allele on the Ph- MPN disease phenotype. Our observations also preclude the use of ABL1 as a housekeeping gene for these neoplasms.
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http://dx.doi.org/10.1007/s00277-013-1920-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3945640PMC
April 2014

Epigenetic role of miRNAs in normal and leukemic hematopoiesis.

Epigenomics 2013 ;5(5):539-52

Department of Medical-Surgical Sciences & Biotechnologies, University La Sapienza, Latina, 04100, Italy.

Hematopoiesis is a regulated multistep process, whereby transcriptional and epigenetic events contribute to progenitor fate determination. miRNAs have emerged as key players in hematopoietic cell development, differentiation and malignant transformation. From embryonic development through to adult life, miRNAs cooperate with, or are regulated, by epigenetic factors. Moreover, recent findings suggest that they contribute to chromatin structural modification, and the functional relevance of this 'epigenetic-miRNA axis' will be discussed in this article. Finally, emerging evidence has highlighted that miRNAs have functional control in human hematopoietic cells, involving targeted recruitment of epigenetic complexes to evolutionarily conserved complementary genomic loci. We propose the existence of epigenetic-miRNA loops that are able to organize the whole gene expression profile in hematopoietic cells.
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http://dx.doi.org/10.2217/epi.13.55DOI Listing
April 2016

Epigenetics in focus: pathogenesis of myelodysplastic syndromes and the role of hypomethylating agents.

Crit Rev Oncol Hematol 2013 Nov 7;88(2):231-45. Epub 2013 Jul 7.

Haematology unit, University of Florence, Florence, Italy. Electronic address:

Dysregulation of cellular epigenetic machinery is considered a major pathogenetic determinant in many malignancies, including myelodysplastic syndromes (MDS). The importance of epigenetic dysfunction in MDS is reflected by the success of hypomethylating agents as standard of care for their treatment. Although these agents improve both survival and quality of life, knowledge gaps remain regarding the precise role of epigenetics in the pathogenesis of MDS and mechanisms by which hypomethylating agents exert their clinical effects. This article reviews the pathogenic role of epigenetic alterations in MDS, including the relationship between genetic and epigenetic abnormalities, and highlights emerging evidence that hypomethylating agents may reprogram the "methylome" while re-establishing hematopoiesis.
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http://dx.doi.org/10.1016/j.critrevonc.2013.06.004DOI Listing
November 2013

Epigenetic silencing of microRNA-193a contributes to leukemogenesis in t(8;21) acute myeloid leukemia by activating the PTEN/PI3K signal pathway.

Blood 2013 Jan 6;121(3):499-509. Epub 2012 Dec 6.

Department of Hematology, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, China.

t(8;21) is one of the most frequent chromosomal translocations occurring in acute myeloid leukemia (AML) and is considered the leukemia-initiating event. The biologic and clinical significance of microRNA dysregulation associated with AML1/ETO expressed in t(8;21) AML is unknown. Here, we show that AML1/ETO triggers the heterochromatic silencing of microRNA-193a (miR-193a) by binding at AML1-binding sites and recruiting chromatin-remodeling enzymes. Suppression of miR-193a expands the oncogenic activity of the fusion protein AML-ETO, because miR-193a represses the expression of multiple target genes, such as AML1/ETO, DNMT3a, HDAC3, KIT, CCND1, and MDM2 directly, and increases PTEN indirectly. Enhanced miR-193a levels induce G(1) arrest, apoptosis, and restore leukemic cell differentiation. Our study identifies miR-193a and PTEN as targets for AML1/ETO and provides evidence that links the epigenetic silencing of tumor suppressor genes miR-193a and PTEN to differentiation block of myeloid precursors. Our results indicated a feedback circuitry involving miR-193a and AML1/ETO/DNMTs/HDACs, cooperating with the PTEN/PI3K signaling pathway and contributing to leukemogenesis in vitro and in vivo, which can be successfully targeted by pharmacologic disruption of the AML1/ETO/DNMTs/HDACs complex or enhancement of miR-193a in t(8;21)-leukemias.
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http://dx.doi.org/10.1182/blood-2012-07-444729DOI Listing
January 2013

Transcriptional targeting by microRNA-polycomb complexes: a novel route in cell fate determination.

Cell Cycle 2012 Oct 16;11(19):3543-9. Epub 2012 Aug 16.

Department of Cellular Biotechnologies and Hematology, University La Sapienza, Rome, Italy.

Advances in the understanding of the epigenetic events underlying the regulation of developmental genes expression and cell lineage commitment are revealing novel regulatory networks. These also involve distinct components of the epigenetic pathways, including chromatin histone modification, DNA methylation, repression by polycomb complexes and microRNAs. Changes in chromatin structure, DNA methylation status and microRNA expression levels represent flexible, reversible and heritable mechanisms for the maintenance of stem cell states and cell fate decisions. We recently provided novel evidence showing that microRNAs, besides determining the post-transcriptional gene silencing of their targets, also bind to evolutionarily conserved complementary genomic seed-matches present on target gene promoters. At these sites, microRNAs can function as a critical interface between chromatin remodeling complexes and the genome for transcriptional gene silencing. Here, we discuss our novel findings supporting a role of the transcriptional chromatin targeting by polycomb-microRNA complexes in lineage fate determination of human hematopoietic cells.
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http://dx.doi.org/10.4161/cc.21468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3478304PMC
October 2012

Polycombs and microRNA-223 regulate human granulopoiesis by transcriptional control of target gene expression.

Blood 2012 Apr 10;119(17):4034-46. Epub 2012 Feb 10.

San Raffaele Bio-medical Park Foundation, Rome, Italy.

Epigenetic modifications regulate developmental genes involved in stem cell identity and lineage choice. NFI-A is a posttranscriptional microRNA-223 (miR-223) target directing human hematopoietic progenitor lineage decision: NFI-A induction or silencing boosts erythropoiesis or granulopoiesis, respectively. Here we show that NFI-A promoter silencing, which allows granulopoiesis, is guaranteed by epigenetic events, including the resolution of opposing chromatin "bivalent domains," hypermethylation, recruitment of polycomb (PcG)-RNAi complexes, and miR-223 promoter targeting activity. During granulopoiesis, miR-223 localizes inside the nucleus and targets the NFI-A promoter region containing PcGs binding sites and miR-223 complementary DNA sequences, evolutionarily conserved in mammalians. Remarkably, both the integrity of the PcGs-RNAi complex and DNA sequences matching the seed region of miR-223 are required to induce NFI-A transcriptional silencing. Moreover, ectopic miR-223 expression in human myeloid progenitors causes heterochromatic repression of NFI-A gene and channels granulopoiesis, whereas its stable knockdown produces the opposite effects. Our findings indicate that, besides the regulation of translation of mRNA targets, endogenous miRs can affect gene expression at the transcriptional level, functioning in a critical interface between chromatin remodeling complexes and the genome to direct fate lineage determination of hematopoietic progenitors.
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http://dx.doi.org/10.1182/blood-2011-08-371344DOI Listing
April 2012

Skeletal muscle regeneration in mice is stimulated by local overexpression of V1a-vasopressin receptor.

Mol Endocrinol 2011 Sep 4;25(9):1661-73. Epub 2011 Aug 4.

Department of Anatomical, Histological, Forensic, and Orthopaedic Sciences, Sapienza University of Rome, Rome, Italy.

Skeletal muscle has a remarkable capacity to regenerate after mechanical or pathological injury. We show that the V1a receptor (V1aR) for vasopressin, a potent myogenic-promoting factor that stimulates differentiation and hypertrophy in vitro, is expressed in mouse skeletal muscle and modulated during regeneration after experimental injury. We used gene delivery by electroporation to overexpress the myc-tagged vasopressin V1aR in specific muscles, thus sensitizing them to circulating vasopressin. The correct localization on the surface of the fibers of the recombinant product was demonstrated by confocal immunofluorescence directed against the myc tag. V1aR overexpression dramatically enhanced regeneration. When compared with mock-transfected controls, V1aR overexpressing muscles exhibited significantly accelerated activation of satellite cells and increased expression of differentiation markers. Downstream of V1aR activation, calcineurin was strongly up-regulated and stimulated the expression of IL-4, a potent mediator of myogenic cell fusion. The central role of calcineurin in mediating V1aR-dependent myogenesis was also demonstrated by using its specific inhibitor, cyclosporine A. This study identifies skeletal muscle as a physiological target of hormones of the vasopressin family and reveals a novel in vivo role for vasopressin-dependent pathways. These findings unveil several steps, along a complex signaling pathway, that may be exploited as potential targets for the therapy of diseases characterized by altered muscle homeostasis and regeneration.
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http://dx.doi.org/10.1210/me.2011-1049DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5417231PMC
September 2011

NFI-A directs the fate of hematopoietic progenitors to the erythroid or granulocytic lineage and controls beta-globin and G-CSF receptor expression.

Blood 2009 Aug 19;114(9):1753-63. Epub 2009 Jun 19.

Department of Histology and Medical Embryology, University La Sapienza, Rome, Italy.

It is generally conceded that selective combinations of transcription factors determine hematopoietic lineage commitment and differentiation. Here we show that in normal human hematopoiesis the transcription factor nuclear factor I-A (NFI-A) exhibits a marked lineage-specific expression pattern: it is upmodulated in the erythroid (E) lineage while fully suppressed in the granulopoietic (G) series. In unilineage E culture of hematopoietic progenitor cells (HPCs), NFI-A overexpression or knockdown accelerates or blocks erythropoiesis, respectively: notably, NFI-A overexpression restores E differentiation in the presence of low or minimal erythropoietin stimulus. Conversely, NFI-A ectopic expression in unilineage G culture induces a sharp inhibition of granulopoiesis. Finally, in bilineage E + G culture, NFI-A overexpression or suppression drives HPCs into the E or G differentiation pathways, respectively. These NFI-A actions are mediated, at least in part, by a dual and opposite transcriptional action: direct binding and activation or repression of the promoters of the beta-globin and G-CSF receptor gene, respectively. Altogether, these results indicate that, in early hematopoiesis, the NFI-A expression level acts as a novel factor channeling HPCs into either the E or G lineage.
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http://dx.doi.org/10.1182/blood-2008-12-196196DOI Listing
August 2009

Epigenetic reprogramming of breast cancer cells by valproic acid occurs regardless of estrogen receptor status.

Int J Biochem Cell Biol 2009 Jan 22;41(1):225-34. Epub 2008 Aug 22.

Department of Histology & Medical Embryology, University of Rome La Sapienza, Italy.

Estrogen receptors (ERs) are a recognized prognostic factor and therapeutic target in breast cancer. The loss of ER expression relates to poor prognosis, poor clinical outcome and impairs the use of anti-estrogenic treatment. Histone deacetylase inhibitors are candidate drugs for cancer therapy. Among them, valproic acid (VPA) is a long used and safe anti-epileptic drug. We studied the biological consequences of the chromatin remodeling action of VPA in a normal human mammary epithelial cell line and in ERalpha-positive and ERalpha-negative breast cancer cell lines. In these cells and regardless of their ER status, VPA-induced cell differentiation, as shown by increased milk lipids production, decreased expression of the CD44 antigen and growth arrest in the G(0)-G(1) phase of the cell cycle. These effects were accompanied by decreased Rb phosphorylation, hyperacetylation of the p21(WAF1/CIP1) gene promoter and increased p21 protein expression. Only in breast cancer cells, cyclin B1 expression was decreased and the cells accumulated also in G(2). ERalpha expression decreased in ERalpha-positive, increased in ERalpha-negative and was unchanged in normal mammary epithelial cells, as did the expression of progesterone receptor, a physiological ERalpha target. VPA decreased the expression of the invasiveness marker pS2 in ERalpha-positive breast cancer cells, but did not cause its re-expression in ERalpha-negative cells. Overall, these data suggest that in both ERalpha-positive and -negative malignant mammary epithelial cells VPA reprograms the cells to a more differentiated and "physiologic" phenotype that may improve the sensitivity to endocrine therapy and/or chemotherapy in breast cancer patients.
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http://dx.doi.org/10.1016/j.biocel.2008.08.019DOI Listing
January 2009

MBD3, a component of the NuRD complex, facilitates chromatin alteration and deposition of epigenetic marks.

Mol Cell Biol 2008 Oct 21;28(19):5912-23. Epub 2008 Jul 21.

Centre de Regulacio Genomica, Universitat Pompeu Fabra, Barcelona, Spain.

In plants, as in mammals, mutations in SNF2-like DNA helicases/ATPases were shown to affect not only chromatin structure but also global methylation patterns, suggesting a potential functional link between chromatin structure and epigenetic marks. The SNF2-like ATPase containing nucleosome remodeling and deacetylase corepressor complex (NuRD) is involved in gene transcriptional repression and chromatin remodeling. We have previously shown that the leukemogenic protein PML-RARa represses target genes through recruitment of DNA methytransferases and Polycomb complex. Here, we demonstrate a direct role of the NuRD complex in aberrant gene repression and transmission of epigenetic repressive marks in acute promyelocytic leukemia (APL). We show that PML-RARa binds and recruits NuRD to target genes, including to the tumor-suppressor gene RARbeta2. In turn, the NuRD complex facilitates Polycomb binding and histone methylation at lysine 27. Retinoic acid treatment, which is often used for patients at the early phase of the disease, reduced the promoter occupancy of the NuRD complex. Knockdown of the NuRD complex in leukemic cells not only prevented histone deacetylation and chromatin compaction but also impaired DNA and histone methylation, as well as stable silencing, thus favoring cellular differentiation. These results unveil an important role for NuRD in the establishment of altered epigenetic marks in APL, demonstrating an essential link between chromatin structure and epigenetics in leukemogenesis that could be exploited for therapeutic intervention.
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http://dx.doi.org/10.1128/MCB.00467-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2546998PMC
October 2008

MicroRNA: basic mechanisms and transcriptional regulatory networks for cell fate determination.

Cardiovasc Res 2008 Sep 6;79(4):553-61. Epub 2008 Jun 6.

Department of Histology and Medical Embryology, 'Sapienza' University of Rome, Italy.

Characterization of regulatory mechanisms affecting microRNA (miRNA) expression and activity is providing novel clues for the identification of genes and complex regulatory circuits that determine cell and tissue specificity. Here, we review the molecular events leading to miRNA biogenesis and activity, focusing above all on endogenous and epigenetic transcriptional networks involving miRNA in early development, cellular lineage specification/differentiation of nervous, skeletal and cardiac muscle tissues and in haematopoiesis, as the de-regulation of such networks may be relevant to disease pathogenesis.
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http://dx.doi.org/10.1093/cvr/cvn151DOI Listing
September 2008

Epigenetic treatment of myelodysplastic syndromes and acute myeloid leukemias.

Curr Med Chem 2008 ;15(13):1274-87

Istituto di Ematologia, Università Cattolica del Sacro Cuore, Rome, Italy.

Epigenetic mechanisms affecting chromatin structure contribute to regulate gene expression and assure the inheritance of information, which are essential for the proper expression of key regulatory genes in healthy cells, tissues and organs. In the medical field, an increasing body of evidence indicates that altered gene expression or de-regulated gene function lead to disease. Cancer cells also suffer a profound change in the genomic methylation patterns and chromatin status. Aberrant DNA methylation patterns, changes in chromatin structure and in gene expression are common in all kind of tumor types. However, studies on leukemias have provided paradigmatic examples for the functional implications of the epigenetic alterations in cancer development and progression as well as their relevance for therapeutical targeting.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2764862PMC
http://dx.doi.org/10.2174/092986708784534947DOI Listing
July 2008

Oncoproteins, heterochromatin silencing and microRNAs: a new link for leukemogenesis.

Epigenetics 2008 Jan-Feb;3(1):1-4

Department of Histology and Medical Embryology; University La Sapienza & San Raffaele Bio-medical Park Foundation; Rome, Italy.

The pathogenesis of acute myeloid leukemias involves complex molecular events triggered by diverse alterations of genomic DNA. A limited number of initiating lesions, such as chromosomal translocations generating fusion genes, are constantly identified in specific forms of leukemia and are critical to leukemogenesis. Leukemia fusion proteins derived from chromosomal translocations can mediate epigenetic silencing of gene expression. Epigenetic deregulation of the DNA methylation status and of the chromatin "histone code" at specific gene sites cooperate in the pathogenesis of leukemias. The neutralization of these crucial oncogenic events can revert the leukemia phenotype. Thus, their identification and the study of their molecular and biological consequences is essential for the development of novel and specific therapeutic strategies. In this context, we recently reported a link between the differentiation block of leukemia and the epigenetic silencing of the microRNA-223 gene by the AML1/ETO oncoprotein, the product of the t(8;21) the commonest AML-associated chromosomal translocation. This finding indicates microRNAs as additional epigenetic targets for leukemogenesis and for therapeutic intervention in leukemias.
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http://dx.doi.org/10.4161/epi.3.1.5651DOI Listing
June 2008

Targeting of the N-terminal coiled coil oligomerization interface by a helix-2 peptide inhibits unmutated and imatinib-resistant BCR/ABL.

Int J Cancer 2008 Jun;122(12):2744-52

Med. Klinik II/Abt. Hämatologie, Johann Wolfgang Goethe-Universität, 60590 Frankfurt, Germany.

The BCR/ABL oncogene is responsible for the phenotype of Philadelphia chromosome-positive (Ph+) leukemia. BCR/ABL exhibits an aberrant ABL-tyrosine kinase activity. The treatment of advanced Ph+ leukemia with selective ABL-kinase inhibitors such as Imatinib, Nilotinib and Dasatinib is initially effective but rapidly followed by resistance mainly because of specific mutations in BCR/ABL. Tetramerization of ABL through the N-terminal coiled-coil region (CC) of BCR is essential for the ABL-kinase activation. Targeting the CC-domain forces BCR/ABL into a monomeric conformation reduces its kinase activity and increases the sensitivity for Imatinib. We show that (i) targeting the tetramerization by a peptide representing the Helix-2 of the CC efficiently reduced the autophosphorylation of both unmutated and mutated BCR/ABL; (ii) Helix-2 inhibited the transformation potential of BCR/ABL independently of the presence of mutations; and (iii) Helix-2 efficiently cooperated with Imatinib as revealed by their effects on the transformation potential and the factor-independence related to BCR/ABL with the exception of mutant T315I. These findings support earlier observations that BCR/ABL harboring the T315I mutation have a transformation potential that is at least partially independent of its kinase activity. These data provide evidence that the inhibition of tetramerization inhibits BCR/ABL-mediated transformation and can contribute to overcome Imatinib-resistance.
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http://dx.doi.org/10.1002/ijc.23467DOI Listing
June 2008

Epigenetic silencing of the myelopoiesis regulator microRNA-223 by the AML1/ETO oncoprotein.

Cancer Cell 2007 Nov;12(5):457-66

Department of Histology and Medical Embryology, University La Sapienza, Rome, 00161, Italy.

Hematopoietic transcription factors are involved in chromosomal translocations, which generate fusion proteins contributing to leukemia pathogenesis. Analysis of patient's primary leukemia blasts revealed that those carrying the t(8;21) generating AML1/ETO, the most common acute myeloid leukemia-associated fusion protein, display low levels of a microRNA-223 (miR-223), a regulator of myelopoiesis. Here, we show that miR-223 is a direct transcriptional target of AML1/ETO. By recruiting chromatin remodeling enzymes at an AML1-binding site on the pre-miR-223 gene, AML1/ETO induces heterochromatic silencing of miR-223. Ectopic miR-223 expression, RNAi against AML1/ETO, or demethylating treatment enhances miR-223 levels and restores cell differentiation. Here, we identify an additional action for a leukemia fusion protein linking the epigenetic silencing of a microRNA locus to the differentiation block of leukemia.
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http://dx.doi.org/10.1016/j.ccr.2007.09.020DOI Listing
November 2007

Dielectric spectroscopy of blood cells suspensions: study on geometrical structure of biological cells.

Conf Proc IEEE Eng Med Biol Soc 2006;2006:3194-7

ICEmB, Rome Univ., Rome, Italy.

A promising procedure combining dielectric spectroscopy of red blood cells and an inverse application of an effective medium theory (EMT) has been realized to prove the plausibility to obtain cells morphological information. This theoretical-experimental methodology could be the basis for an accurate and simple tool in diagnosis and research activity, especially when cell morphological alterations are a prime indicator of illness as in all hematopoietic pathologies.
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http://dx.doi.org/10.1109/IEMBS.2006.259414DOI Listing
March 2008