Irene Filippi

Università degli Studi di Siena, Siena, Tuscany, Italy

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Publications (19)63.88 Total impact

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    Irene Filippi · Fabio Carraro · Antonella Naldini
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    ABSTRACT: Inflammation and tumor hypoxia are intimately linked and breast cancer provides a typical example of an inflammation-linked malignant disease. Indeed, breast cancer progression is actively supported by inflammatory components, including IL-1 β , and by the hypoxia-inducible factor- (HIF-) 1 α . In spite of many attempts where the role of either IL-1 β or HIF-1 α was evaluated, detailed mechanisms for their effects on breast cancer cell migration under hypoxia are still unclear. We here report that IL-1 β increased MDAMB231 cell migration under hypoxic conditions along with HIF-1 α accumulation and upregulation of CXCR1, which is transcriptionally regulated by HIF-1 α , as well as an increased expression of CXCL8 and NF κ B. In addition, IL-1 β -induced cell migration in hypoxia was not affected when HIF-1 α was inhibited by either siRNA or Topotecan, well known for its inhibitory effect on HIF-1 α . Of interest, HIF-1 α inhibition did not reduce NF κ B and CXCL8 expression and the reduction of IL-1 β -induced cell migration under hypoxia was achieved only by pharmacological inhibition of NF κ B. Our findings indicate that inhibition of HIF-1 α does not prevent the migratory program activated by IL-1 β in hypoxic MDAMB231 cells. They also suggest a potential compensatory role of NF κ B/CXCL8 pathway in IL-1 β -induced MDAMB231 cell migration in a hypoxic microenvironment.
    Full-text · Article · Nov 2015 · Mediators of Inflammation

  • No preview · Conference Paper · Oct 2015
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    ABSTRACT: Endometriosis is associated with local angiogenic and hypoxic mechanisms. Indeed, peritoneal fluid of women with endometriosis generates a specific microenvironment to support the growth and development of ectopic endometrial tissues. The association between proangiogenic markers and hypoxic processes in different endometriosis phenotypes was investigated in the present study, analyzing the expression of several genes, related to hypoxic signaling pathway and involved in angiogenic processes, in nonpregnant women with different forms of endometriosis. Samples of ovarian endometrioma (OMA; n = 16) or deep infiltrating endometriosis (DIE; n = 11) were collected, and in addition, control endometrium was collected from healthy women by hysteroscopy. The gene expression of the hypoxia-inducible factors (HIF) 1/2α, protease-activated receptors (PARs) ¼, and vascular endothelial growth factor (VEGF) A was evaluated by quantitative reverse-transcription polymerase chain reaction. Ovarian endometrioma expresses high levels of HIF-1/2α, PAR-1/4, and VEGF-A, while DIE did not show significantly different gene expression compared to endometrium from unaffected women. A positive correlation between the expression of HIF-1/2α and VEGF-A mRNA was observed in OMA. The overall data point out that the heterogeneity of the disease reflects differences in expression levels of genes associated with hypoxia and angiogenesis, suggesting that such conditions may have an active role in the development of the disease.
    No preview · Article · Sep 2015 · Reproductive sciences (Thousand Oaks, Calif.)
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    ABSTRACT: Hypoxia represents an inadequate oxygen supply to tissues, which can modulate cell functions, primarily through the hypoxia-inducible transcription factor HIF-1α. Dendritic cells (DC) are professional antigen-presenting cells and their migration maybe affected by hypoxia, since the local microenvironment in lymphoid organs, as well as in inflamed and tumor tissues, is characterized by low oxygen tensions. In this study we observed an enhanced migratory capability of human monocyte-derived DC, using in vitro migration assays performed under hypoxic conditions. Such enhancement was independent on either the chemoattractants involved or the maturation level of DC. However, HIF-1α appeared to be crucial for the migration only of immature DC and not for mature DC under hypoxia, as indicated by HIF-1α siRNA approaches. Furthermore, we observed that while Akt phosphorylation was enhanced in both immature and mature DC exposed to hypoxia, other signaling pathways, such as p38 and p42/p44 MAPK, were differently affected during hypoxic treatment. More interestingly, aspecific and specific inhibition of PI3K/Akt indicated that such pathway was relevant for the migration of both immature and matured DC under hypoxia, even when DC were transfected with HIF-1α siRNA. Our results indicate that, besides HIF-1α, several other pathways, including PI3K/Akt, may be involved in the response to hypoxia of immature and, more specifically, of mature DC to sustain their trafficking and functions within hypoxic microenvironments. J. Cell. Physiol. 9999: XX–XX, 2014. © 2014 Wiley Periodicals, Inc.
    No preview · Article · Dec 2014 · Journal of Cellular Physiology
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    ABSTRACT: Starting from our in-house library of pyrazolo[3,4-d]pyrimidines, cross-docking simulation was conducted on Bcr-Abl T315I mutant to select new derivatives for biological investigations. Among the selected compounds (2a-e), derivative 2b showed a high activity against the Bcr-Abl T315I mutant (Ki = 36 nM). Binding free energy calculation (MM-GBSA), molecular interaction field (MIF) analysis and free energy perturbation (FEP) studies highlighted the importance of a bromine atom of the para position of the N1 side chain phenyl ring for the interaction with the hydrophobic region I in the T315I mutant. A series of 4-bromo derivatives was thus synthesized and biologically evaluated in cell-free assays (c-Src, Abl wt, Abl T315I mutant) and in the murine myeloid 32D cell lines expressing the human wild type p210-Bcr-Abl or the Bcr-Abl T315I mutant. Compound 2j was identified as the most promising derivative showing a good balance of different ADME properties, high activity in cell-free assays and an interesting sub-micromolar potency against T315I Bcr-Abl expressing cells. In addition, liposome encapsulated 2j was tested on 32D-p210 and 32D-T315I cell lines at concentrations of 0.1 and 1 μM in comparison with the DMSO dissolved 2j. Liposomal formulation increases the solubility of pyrazolo[3,4-d]pyrimidines preserving a good activity on leukemic T315I cells and avoiding the use of DMSO as solubilizing agent. In vivo studies on mice inoculated with 32D-T315I cells and treated for 17 days with 2j showed a more than 50% reduction in tumor volumes when compared to placebo treated mice.
    No preview · Article · Jun 2013 · Journal of Medicinal Chemistry
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    ABSTRACT: N-[2-Methyl-5-(triazol-1-yl)phenyl]pyrimidin-2-amine derivatives were synthesized and evaluated in vitro for their potential use as inhibitors of Bcr-Abl. The design is based on the bioisosterism between the 1,2,3-triazole ring and the amide group. The synthesis involves a copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) as the key step, with the exclusive production of anti-(1,4)-triazole derivatives. One of the compounds obtained shows general activity similar to that of imatinib; in particular, it was observed to be more effective in decreasing the fundamental function of cdc25A phosphatases in the K-562 cell line.
    Full-text · Article · Nov 2011 · ChemMedChem
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    ABSTRACT: The tumor microenvironment is characterized by a poor circulation which results in the selection of neoplastic cells that can grow or survive under hypoxic conditions. The relationship between hypoxia and histone deacetylase (HDAC) inhibitors has been previously established. In this work we evaluated the effects of novel HDAC inhibitors (the natural peptide FR235222 and three tetrapeptide analogs) in the human breast cancer cell line MDAMB231, cultured under hypoxia (2% O2 ≉ 14 mmHg) or normoxia (20% O2 ≉ 140 mmHg). First, we found that the novel HDAC inhibitors reduced cell proliferation in MDAMB231 cells at an extent which was similar or even higher than that exerted by the classic HDAC inhibitors trichostatin-A and suberoylanilide hydroxamic acid. More interestingly, the antiproliferative effects of the novel HDAC inhibitors were, in general, significantly higher in hypoxic cells than in normoxic controls. Hypoxic MDAMB231 cells expressed high levels of the hypoxia-inducible factor (HIF)-1α and HIF-1α-related genes, such as vascular endothelial growth factor, Bcl-2/E1B 19 kDa interacting protein-3, glucose transporter-1, carbonic anhydrase IX, as determined by Western blot analysis and qRT-PCR. Finally, we found that HIF-1α and HIF-1α-related genes were significantly downregulated by FR235222 and analogs. In conclusion, the identification of novel effects exerted by the HDAC inhibitors, characterized by a strong efficacy in inhibiting the expression of HIF-1α and its related genes, may have important implications in the pharmacological control of several tumors, including breast cancer, characterized by the presence of hypoxia, angiogenesis and metabolic derangements.
    Full-text · Article · Oct 2011 · Anti-cancer agents in medicinal chemistry

  • No preview · Conference Paper · Aug 2011
  • I Filippi · A Naldini · F Carraro
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    ABSTRACT: The tumor microenvironment is characterized, not only by marked gradients in drug concentration, but also by gradients in the rate of cell proliferation and by regions of hypoxia and acidity, all of which can influence tumor cell sensitivity to drug treatment. Hypoxia is also an important environmental factor in chronic myeloid leukemia (CML), because bone marrow is intrinsically hypoxic in nature. Systems-wide analyses of tumors have recently identified receptor tyrosine kinase coactivation as an important mechanism by which cancer cells achieve chemoresistance. Recent work suggests that Src activation might play a prominent role in the response to hypoxia to promote cell survival, progression, and metastasis of a variety of human cancer. Other studies also established a functional link between Bcr-Abl and the Src family tyrosine kinases. It is well known that mutations can also cause some tyrosine kinases to become constitutively active, a nonstop functional state that may contribute to initiation or progression of cancer as in CML. Leukemic cells carrying chromosomal alteration, are sensitive to imatinib that induces complete remission in most patients. This inhibitor is a highly selective Bcr-Abl tyrosine kinase inhibitor (TKI). There is a considerable interest in understanding how activated signaling pathways enhance tumor cell survival under hypoxia, because this might lead to the introduction of more effective treatments to target these resistant subpopulations. For all these reasons it is important to identify new TKIs which are also active in hypoxia, the real tumor microenvironment, as possible alternative therapy.
    No preview · Article · Jun 2011 · Current Medicinal Chemistry
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    ABSTRACT: A family of dual Src/Abl inhibitors characterized by a substituted pyrazolo[3,4-d]pyrimidine scaffold was previously reported by us and proved to be active against several tumor cell lines. Among these compounds, a promising antileukemia lead (1) has been recently identified, but, unfortunately, it suffers from substandard pharmaceutical properties. Accordingly, an approach for the optimization of the lead 1 is described in the present work. A series of more soluble pyrazolo[3,4-d]pyrimidine derivatives were rationally designed and proved to maintain the dual Src/Abl activity of the lead. Selected compounds showed an interesting activity profile against three different leukemic cells also in hypoxic conditions, which are usually characterized by imatinib-resistance. Finally, in vitro ADME properties (PAMPA permeation, water solubility, microsomal stability) for the most promising inhibitors were also evaluated, thus allowing the identification of a few optimized analogues of lead 1 as promising antileukemia agents.
    No preview · Article · Mar 2011 · Journal of Medicinal Chemistry
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    ABSTRACT: The casual relationship between inflammation and tumour progression has been widely accepted and the etiology of breast cancer has been associated with inflammatory processes. Interleukin (IL)-1β, besides its central role in inflammation, has also been recognised as a powerful player in tumour progression, angiogenesis and invasiveness. Recently, there has been considerable interest in understanding the non-hypoxic upregulation of the hypoxia-inducible factor (HIF)-1α by IL-1 in neoplastic cells since aberrant expression of HIF-1α correlates with tumour progression. Here, using the highly invasive human breast cancer cell line MDAMB231, we studied the effect of IL-1β on tumour cell migration along with HIF-1α accumulation. We observed that non-hypoxic induction of HIF-1α by IL-1β in MDAMB231 was associated with increased cell migration, paralleled by upregulation of p38 MAPK phosphorylation and CXCL8/CXCR1 expression. Inhibition of HIF-1α by siRNA resulted in a significant reduction of CXCR1 expression and IL-1β-induced cell migration in MDAMB231 cells, thus confirming a role of HIF-1α in the non-hypoxic-IL-1β-dependent induction of migratory potentials. Our observation that IL-1 induces HIF-1α accumulation in MDAMB231 cells was confirmed in tumour cells growing in vivo using an experimental approach, mimicking the endogenous release of IL-1 in mice bearing MDAMB231 xenografts. Our in vivo data, along with the fact that inhibition of HIF-1α resulted in the decrease of IL-1β-promoted cell migration, further support the link between inflammation and cancer. The overall results may have important implications in those therapeutic approaches aimed to inhibit IL-1-mediated activities in tumour cells, specifically in breast cancer.
    No preview · Article · Dec 2010 · European journal of cancer (Oxford, England: 1990)
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    ABSTRACT: Potential boron neutron capture therapy (BNCT) agents have been designed on the basis of the evidence about translocator protein (TSPO) overexpression on the outer mitochondrial membrane of tumor cells. The structure of the first TSPO ligand bearing a carborane cage (compound 2d) has been modified in order to find a suitable candidate for in vivo studies. The designed compounds were synthesized and evaluated for their potential interaction with TSPO and tumor cells. In vitro biological evaluation showed in the case of fluoromethyl derivative 4b a nanomolar TSPO affinity very similar to that of 2d, a significantly lower cytotoxicity, and a slightly superior performance as boron carrier toward breast cancer cells. Moreover, compound 4b could be used as a ¹⁹F magnetic resonance imaging (MRI) agent as well as labeled with ¹¹C or ¹⁸F to obtain positron emission tomography (PET) radiotracers in order to apply the "see and treat" strategy in BNCT.
    No preview · Article · Nov 2010 · Bioconjugate Chemistry
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    ABSTRACT: The main aim of this study was to enhance the solubility of pyrazolo[3,4-d]pyrimidines 1-8 able to strongly inhibit Src and Abl tyrosine kinase phosphorylation in cell-free assays and to significantly reduce leukemic and osteosarcoma cell lines growth, but characterized by very low solubility in aqueous media. Their water solubility was improved between 100 and 1000 folds by solubilization with 2-hydroxypropyl-β-cyclodextrin (HPβCD) and ratio of inclusion complex were determined by phase solubility method. Finally, some complexed compounds were tested on different leukemic (K-652, KU-812 and HL-60) and osteosarcoma (SaOS-2) cell lines showing a good enhancement of biological response in comparison with the not complexed compounds.
    No preview · Article · Oct 2010 · European Journal of Medicinal Chemistry

  • No preview · Article · Apr 2009 · Pigment Cell & Melanoma Research
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    ABSTRACT: Aberrant expression of the protease-activated receptor (PAR)-1 has been associated with tumour progression. Indeed, PAR-1 expression correlates with tumour invasiveness, as well as with cancer cell survival. As the tumour microenvironment is characterised by a low oxygen tension, we decided to investigate the role of PAR-1 in cancer cells exposed to a hypoxic microenvironment. In this study we show that hypoxia enhances PAR-1 expression in MDAMB231 breast cancer cells. We next provided evidence for a novel role of PAR-1 in protecting hypoxic breast cancer against cell death, since inhibition of PAR-1 by RNA interference resulted in a decreased cell survival. Finally, we found that treatment of hypoxic MDAMB231 cells with the specific PAR-1 agonist peptide (TRAP) resulted in a significant increase of cell survival and migration. The overall results identify for the first time a functional role for PAR-1 in the cellular responses of breast cancer to a hypoxic microenvironment.
    Full-text · Article · Dec 2008 · European journal of cancer (Oxford, England: 1990)
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    ABSTRACT: The ability to maintain O(2) homeostasis is essential to the survival of all invertebrate and vertebrate species. The transcriptional factor, hypoxia inducible factor 1 (HIF-1), is the principal regulator of oxygen homeostasis. Under hypoxic condition HIF-1 induces the transcription of several hypoxia-responsive genes by binding to hypoxia-response elements (HRE) in their promoters. In recent years it has been demonstrated that hypoxia could be related to metabolic variations such as hyper-cholesterolemia in mouse models. On the basis of this observation, the present study was performed to verify the involvement of HIF-1, and in particular the effect of chemical and environmental induction of HIF-1alpha (the oxygen sensitive isoform) accumulation in 3-hydroxy 3-methylglutaryl coenzyme A reductase (HMG-CoAR, the key and rate limiting enzyme of cholesterol biosynthetic pathway) regulation. Our results show that HIF-1alpha accumulation is able to increase level and activity of HMG-CoAR by stimulating its transcription. The raised transcription of the reductase could be related to an induction by HIF-1alpha even if a parallel action of SREBP-2 actively translocated to nucleus by the increased level of SCAP cannot be excluded.
    Full-text · Article · Jun 2008 · Journal of Cellular Biochemistry
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    ABSTRACT: The ability to maintain O 2 homeostasis is essential to the survival of all invertebrate and vertebrate species. The transcriptional factor, hypoxia inducible factor 1 (HIF-1), is the principal regulator of oxygen homeostasis. Under hypoxic condition HIF-1 induces the transcription of several hypoxia-responsive genes by binding to hypoxia-response elements (HRE) in their promoters. In recent years it has been demonstrated that hypoxia could be related to metabolic variations such as hyper-cholesterolemia in mouse models. On the basis of this observation, the present study was performed to verify the involvement of HIF-1, and in particular the effect of chemical and environmental induction of HIF-1a (the oxygen sensitive isoform) accumulation in 3-hydroxy 3-methylglutaryl coenzyme A reductase (HMG-CoAR, the key and rate limiting enzyme of cholesterol biosynthetic pathway) regulation. Our results show that HIF-1a accumulation is able to increase level and activity of HMG-CoAR by stimulating its transcription. The raised transcription of the reductase could be related to an induction by HIF-1a even if a parallel action of SREBP-2 actively translocated to nucleus by the increased level of SCAP cannot be excluded.
    Full-text · Dataset · Jan 2008
  • I. Filippi · C. Ardinghi · R. Giavazzi · F. Carraro · A. Naldini

    No preview · Article · Dec 2007 · Thrombosis Research
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    ABSTRACT: Thrombin, the key enzyme of the coagulation cascade, is involved in inflammation. It was proposed recently that thrombin activity may play an important role in allergic inflammation. Interferon-gamma (IFN-gamma) is a potent Th1-related cytokine secreted by activated T cells and is usually downregulated in allergic inflammation. We recently demonstrated that thrombin enhances interleukin-10 (IL-10) in peripheral blood mononuclear cells (PBMC). Thus, we hypothesized that thrombin may promote a Th2 profile. We here report that human alpha- thrombin downregulates IFN-gamma expression at both protein and mRNA levels in activated PBMCs. The use of proteolytically inactive thrombin and of the specific thrombin receptor agonist peptide, SFLLRN, shows that this downregulation is thrombin specific and requires thrombin proteolytic activity. The addition of an anti- IL-10 monoclonal antibody (mAb) to thrombin-treated PBMCs abolishes IFN-gamma downregulation, suggesting that thrombin exerts its effect through IL-10, a Th2-related cytokine. Furthermore, IFN-gamma reduction was accompanied by increased IL-4 release, as well as by an increase in the proinflammatory cytokine IL-1. In conclusion, the observation that thrombin affects the production of IFN-gamma (Th1 profile) and IL-4 (Th2 profile) provides further evidence for the role played by thrombin in modulating Th1/Th2 cytokine balance, which could be particularly relevant in allergic inflammation.
    No preview · Article · Dec 2006 · Journal of Interferon & Cytokine Research