92 research outputs found
Spectroscopic/Computational Characterization and the X-ray Structure of the Adduct of the VIVO-Picolinato Complex with RNase A
No full textThe structure, stability, and enzymatic activity of the adduct formed upon the reaction of the V-picolinato (pic) complex [VIVO(pic)2(H2O)], with an octahedral geometry and the water ligand in cis to the V=O group, with the bovine pancreatic ribonuclease (RNase A) were studied. While electrospray ionization-mass spectrometry, circular dichroism, and ultraviolet-visible absorption spectroscopy substantiate the interaction between the metal moiety and RNase A, electron paramagnetic resonance (EPR) allows us to determine that a carboxylate group, stemming from Asp or Glu residues, and imidazole nitrogen from His residues are involved in the V binding at acidic and physiological pH, respectively. Crystallographic data demonstrate that the VIVO(pic)2 moiety coordinates the side chain of Glu111 of RNase A, by substituting the equatorial water molecule at acidic pH. Computational methods confirm that Glu111 is the most affine residue and interacts favorably with the OC-6-23-Δenantiomer establishing an extended network of hydrogen bonds and van der Waals stabilizations. By increasing the pH around neutrality, with the deprotonation of histidine side chains, the binding of the V complex to His105 and His119 could occur, with that to His105 which should be preferred when compared to that to the catalytically important His119. The binding of the V compound affects the enzymatic activity of RNase A, but it does not alter its overall structure and stability
Enhanced 5-FU-mediated activation of p53-dependent nucleolar stress by MDM2/4 peptide inhibitors in colon cancer cells
No full textRecent evidence suggest that the p53-dependent nucleolar stress induced by some anticancer drugs, such as 5-Fluorouracil (5-FU), might not be sufficient to produce cell cycle arrest and/or apoptosis, given the hyperactivation of MDM2 and MDM4, main inhibitors of the p53 activity. Besides, it has been recently discovered that the disruption of the MDM2/4 heterodimer produce a more efficient reactivation of p53. Pep3 is a 12-mer peptide able to reduce p53 ubiquitination, representing the only peptide inhibitor of the MDM2/4 complex known to date. In this study, we developed a library of Pep3 derivatives with sequence lengths ranging from 5- to 8-mer. Of note, the newly truncated peptides showed lower IC50 values than Pep3. We incorporated the most active peptides (VLP-13 and VLP-24), alone or in association with 5-FU, in biodegradable nanoparticles, and we analyzed their cytotoxicity against p53 proficient colon cancer cells. We found that the cytotoxic activity of VLP-24 was higher than VLP-13 both alone and in association with 5-FU. The combined treatment 5-FU/VLP-24 caused a significant unbalance of ribosomal RNA precursor levels associated to a strong increase of p53 expression levels triggering apoptosis. In conclusion, our data reveal VLP-24 as a promising molecule able to enhance 5-FU mediated activation of p53-dependent nucleolar stress
MDM2/4 inhibitors as novel therapeutic strategy to enhance 5-FU induced nucleolar stress and apoptosis in colon cancer cells
No full textCombination of 5-FU and MDM2/4 peptide inhibitors as a promising strategy to enhance 5-FU induced nucleolar stress.
No full textThe inactivation of tumor suppressor p53 is crucial for tumor onset and progression. Half of colorectal cancer cases carry wild-type p53 that is inactivated by the overexpression of MDM2 and MDM4, the main negative regulators of p53 activity [1]. Hence, the reactivation of p53 by inhibiting MDM2/4-p53 interaction represents a useful strategy to enhance the p53-depemdent cancer cell response to chemotherapeutic drugs. Recently, it has been demonstrated that 5-Fluoruracil (5-FU), the standard first-line treatment for colorectal cancer therapy, exerts its cytotoxic effect through the activation of p53-dependent nucleolar stress response [2].
It has been recently discovered that Pep3, a 12-mer peptide, is able to disrupt the MDM2/4 heterodimer producing a more efficient p53 reactivation [3]. In this study, we developed a library of sixteen Pep3 derivatives with sequence lengths ranging from 5- to 8-mer. Intriguingly, the newly truncated peptides displayed lower IC50 values than Pep3; among which the most active peptides were VLP-13 and VLP-24. We incorporated them, alone or in association with 5-FU, in novel biodegradable nanoparticles, and we analyzed their cytotoxic activity against p53 proficient colon cancer cells. We found that the cytotoxic activity of VLP-24 was higher than VLP-13 alone or in association with 5-FU. Notably, the combined treatment with 5-FU and VLP-24 caused a significant unbalance of ribosomal RNA precursor levels associated to a strong increase of p53 and p21 expression levels leading to the activation of apoptosis. In conclusion, our data demonstrate that VLP-24 is a promising strategy to enhance 5-FU mediated activation of p53-dependent nuclear stress response.
[1] Liebl MC et al. Cancers (2021); 13, 2125.
[2] Pecoraro A et al. Int J Mol Sci. (2021); 22, 5496.
[3] Pellegrino M et al. Cancer Res. (2015); 75; 4560-72
State of the Art on the European Court of Justice and Enacting Citizenship. CEPS Special Report, April 27 2009
No full textThis report provides a state of the art of the main interdisciplinary academic discussions, EU acts and European Court of Justice (ECJ) case law surrounding issues related to citizenship, migration and integration. The report was finalised in mid-2008 and has provided the basis upon which the work conducted by the Justice and Home Affairs Section at CEPS in the framework of the ENACT research project funded by DG Research of the European Commission has been developed. In particular, the general objectives of CEPS’ contribution to this project are: first, to assess the impact of Community governance on the enactment of European citizenship and the exclusivity of the nation-state competence over nationality matters; and to examine the ways in which the ECJ and the adoption of the Council Directive 2004/38/EC on the rights of citizens of the Union and their family members to move and reside freely have influenced its enactment; second, to analyse the impacts of the enlargement processes, and of accompanying measures such as the transitional arrangements inserted in the Acts of Accession and other restrictions to the fundamental right of freedom of movement, on the status and practices of European citizenship; and third, to assess the tensions inherent to nationality and/or residence-based enactment of citizenship versus European citizenship of TCNs; to address the effects and dilemmas posed by the Council Directive 2003/109/EC of November 2003 on the status of third country nationals who are long-term residents
Comparative Analysis of the Inhibitory Mechanism of Aβ1–42 Aggregation by Diruthenium Complexes
No full textThere is a growing interest in the search for metal-based therapeutics for protein misfolding disorders such as Alzheimer’s disease (AD). A novel and largely unexplored class of metallodrugs is constituted by paddlewheel diruthenium complexes, which exhibit unusual water solubility and stability and unique coordination modes to proteins. Here, we investigate the ability of the complexes [Ru2Cl(DPhF)(O2CCH3)3]·H2O (1), [Ru2Cl(DPhF)2(O2CCH3)2]·H2O (2), and K2[Ru2(DPhF)(CO3)3]·3H2O (3) (DPhF– = N,N′-diphenylformamidinate) to interfere with the amyloid aggregation of the Aβ1–42 peptide. These compounds differ in charge and steric hindrance due to the coordination of a different number of bulky ligands. The mechanisms of action of the three complexes were studied by employing a plethora of physicochemical and biophysical techniques as well as cellular assays. All these studies converge on different mechanisms of inhibition of amyloid fibrillation: complexes 1 and 2 show a clear inhibitory effect due to an exchange ligand process in the Ru2 unit aided by aromatic interactions. Complex 3 shows no inhibition of aggregation, probably due to its negative charge in solution. This study demonstrates that slight variations in the ligands surrounding the bimetallic core can modulate the amyloid aggregation inhibition and supports the use of paddlewheel diruthenium complexes as promising therapeutics for Alzheimer’s disease.Universidad Complutense de MadridDepto. de Química InorgánicaFac. de Ciencias QuímicasTRUEpu
A novel ErbB2 epitope targeted by Human antitumor Immunoagents
No full textWe have engineered novel fully human anti-ErbB2 immunoagents: Erbicin, a human scFv (1); ERB-hRNase, a human immunoRNase made up of Erbicin fused to a human RNase (2); ERB-hcAb, a human, “compact” antibody, in which two Erbicin molecules are fused to the Fc of a human IgG1(3,4). Both ERB-hRNase and ERB-hcAb severely inhibit the growth of ErbB2-positive tumours sensitive and resistant to Trastuzumab treatment (5). Furthermore, they are not immunogenic and do not display cardiotoxic effects in vitro and in vivo whereas Trastuzumab was strongly toxic (6).
An analysis by three independent methodologies of the interactions of Erbicin-Derived-ImmunoAgents (EDIAs) and Trastuzumab with the ErbB2 extracellular domain (ECD) led to the finding that EDIAs bind soluble ECD with a lower affinity than that for the native receptor on tumor cells. Trastuzumab instead shows a higher affinity for soluble ErbB2-ECD.
Accordingly, ErbB2-ECD abolished the biological properties of Trastuzumab with no effects on those of EDIAs (7). These results suggest that the fraction of immunoagent neutralized by free ECD shed into the bloodstream is much higher for Trastuzumab than for EDIAs, thus the latter could be used at lower therapeutic doses compared to those employed with Trastuzumab.
More interestingly, we found through the use of three different methods, that the epitope recognized by EDIAs is different from those recognized by Trastuzumab and Pertuzumab, and it is located in the domain I of the extracellular region of ErbB2. This finding could lead to the identification of novel epitopes on ErbB2 to be used as potential therapeutic targets to mitigate anti-ErbB2 associated cardiotoxicity and eventually overcome resistance.
1De Lorenzo, C.; Palmer, DB.; Piccoli, R; Ritter, MA.; and D'Alessio, G.; Clin. Cancer Res. 2002, 8, 1710-1719.
2De Lorenzo, C.; Arciello, A.; Cozzolino, R.; Palmer, DB.; Laccetti, P.; Piccoli, R; and D'Alessio, G.; Cancer Res. 2004, 64, 4870-4874.
3De Lorenzo, C.; Tedesco, A.; Terrazzano, G.; Cozzolino, R.; Laccetti, P.; Piccoli, R.; and D'Alessio, G.; Br. J. Cancer 2004, 91, 1200-1204.
4De Lorenzo, C.; Cozzolino, R.; Carpentieri, A.; Pucci, P.; Laccetti, P.; and D'Alessio, G.; Carcinogenesis 2005, 26 ,1890-1895.
5 Gelardi, T.; Damiano, V.; Rosa, R..; Bianco, R..; Cozzolino, R.; Tortora, G.; Laccetti, P.; D'Alessio, G.; and De Lorenzo, C; Br. J. Cancer 2010, 102, 513-519.
6 Riccio, G.; Esposito, G.; Leoncini, E.; Contu, R..; Condorelli, G.; Chiariello, M.; Laccetti, P.; Hrelia, S.; D'Alessio, G.; and De Lorenzo, C.; Faseb J. 2009, 23, 3171-3178
7Troise, F.; Cafaro,V.; Giancola, C.; D’Alessio G.; and De Lorenzo, C.; FEBS J. 2008, 275, 4967–4979
Heterobimetallic ferrocenyl-chromone compounds as selective inhibitors of amyloid aggregation
No full textAmyloid aggregation is a key process in neurodegeneration, producing toxic species that contribute to disease progression. This underscores the urgent need to identify novel agents capable of reducing toxicity by modulating this aggregation process. Two heterobimetallic complexes incorporating a ferrocenyl-chromone (Fc-Chr) core were investigated: one featuring an additional gold(I) triphenylphosphine moiety, Fc-Chr-AuP(Ph)3, and the other containing a dicobalt hexacarbonyl-alkyne unit, Fc-Chr-Co2(CO)6. Their effects were evaluated toward on the aggregation of two amyloid models: the peptide spanning residues 264–277 of nucleophosmin 1 (NPM1264–277) and the C-terminal fragment of the amyloid-β peptide (Aβ21–40) each with unique primary sequences, self-aggregation mechanisms and kinetics. Thioflavin T- assays allowed to assess the impact of the metal complexes on the aggregation of two amyloids. Results indicate that the two complexes inhibit the early stages of the aggregation of peptides in a dose-dependent manner and a greater effect on NPM1264–277 when compared to Aβ21–40 was observed. Native electrospray ionization mass spectrometry revealed the formation of peculiar metal/peptide adducts in dependence on different metal-units in the complexes. Scanning electron microscopy (SEM) and density function theory (DFT) were also employed to further characterize the interaction between the metal compounds and the investigated peptides, while preliminary cell viability assays in SH-SY5Y cells supported inhibitory effects on the aggregation and showed reduction of amyloid cytotoxicity. Fc-Chr-Co2(CO)6 demonstrated the highest efficacy in modulating peptide aggregation, exerting a more significant impact on NMP1264–277 relative to Aβ21−40. These results support the use of ferrocenyl-chromone containing metal complexes as modulators of amyloid peptide aggregation
A Diruthenium Metallodrug as a Potent Inhibitor of Amyloid-β Aggregation: Synergism of Mechanisms of Action
No full text: The physical and chemical properties of paddlewheel diruthenium compounds are highly dependent on the nature of the ligands surrounding the bimetallic core. Herein, we compare the ability of two diruthenium compounds, [Ru2Cl(D-p-FPhF)(O2CCH3)3]·H2O (1) (D-p-FPhF- = N,N'-bis(4-fluorophenyl)formamidinate) and K3[Ru2(O2CO)4]·3H2O (2), to act as inhibitors of amyloid aggregation of the Aβ1-42 peptide and its peculiar fragments, Aβ1-16 and Aβ21-40. A wide range of biophysical techniques has been used to determine the inhibition capacity against aggregation and the possible mechanism of action of these compounds (Thioflavin T fluorescence and autofluorescence assays, UV-vis absorption spectroscopy, circular dichroism, nuclear magnetic resonance, mass spectrometry, and electron scanning microscopy). Data show that the most effective inhibitory effect is shown for compound 1. This compound inhibits fiber formation and completely abolishes the cytotoxicity of Aβ1-42. The antiaggregatory capacity of this complex can be explained by a binding mechanism of the dimetallic units to the peptide chain along with π-π interactions between the formamidinate ligand and the aromatic side chains. The results suggest the potential use of paddlewheel diruthenium complexes as neurodrugs and confirm the importance of the steric and charge effects on the properties of diruthenium compounds
Musculoskeletal disorders and diseases in healthcare workers. A scoping review
No full textBackground: The impact of work-related tasks with potential biomechanical overload on the musculoskeletal system represents an essential socio-economical challenge. Objective: This scoping review aimed to map the current literature to identify musculoskeletal disorders and diseases among healthcare professionals and define the approaches used to assess these problems. Methods: The analysis was developed according to the JBI methodologies for scoping reviews and reported following the PRISMA-ScR framework. Results: We conducted specific searches on online databases; of the 357 articles initially identified, only 118 met the inclusion criteria. Conclusions: Despite some discrepancies in their unambiguous identification, the presence of work-related musculoskeletal disorders in healthcare professionals is unequivocal. On the other hand, using technology as a supporting evaluation tool still needs to be explored. Furthermore, several improvements are required to enhance the quality of work and simplify the analysis across studies
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