Analytical Biochemistry
Biomarker discovery
The research team has gathered considerable experience in the field of biomarker discovery trough proteomics and metabolomics approaches in multifactorial diseases (Figure 1), with a focus on Multiple Sclerosis (MS). To achieve this goal many milestone papers on samples handling has been published, describing molecular alterations in serum and cerebrospinal fluid (CSF) samples from MS patients. In particular, through a top-down proteomics approach, we identified post-translational modifications of CSF transthyretin specifically associated to MS. However, difficult sampling limits the use of CSF biomarkers in large-scale screenings, thus our group is switching its focus on tears, an easily accessible biological fluid which may open a novel window on the nervous system. The quantification of proteins, lipids, metabolites and hormones in tears is also proving useful for the identification of markers related to several other diseases.
Cicalini I, Rossi C, Pieragostino D, Agnifili L, Mastropasqua L, di Ioia M, De Luca G, Onofrj M, Federici L, Del Boccio P. Integrated Lipidomics and Metabolomics Analysis of Tears in Multiple Sclerosis: An Insight into Diagnostic Potential of Lacrimal Fluid. Int J Mol Sci. 2019 Mar 13;20(6). pii: E1265. doi: 10.3390/ijms20061265. Rossi C, Cicalini I, Zucchelli M, di Ioia M, Onofrj M, Federici L, Del Boccio P, Pieragostino D. Metabolomic Signature in Sera of Multiple Sclerosis Patients during Pregnancy. Int J Mol Sci. 2018 Nov 14;19(11). pii: E3589. doi: 10.3390/ijms19113589. Pieragostino D*, Agnifili L, Cicalini I, Calienno R, Zucchelli M, Mastropasqua L, Sacchetta P, Del Boccio P, Rossi C. M. Tear Film Steroid Profiling in Dry Eye Disease by Liquid Chromatography Tandem Mass Spectrometry. Int. J. Mol. Sci. 2017, 18(7), 1349; doi:10.3390/ijms18071349. Del Boccio P, Perrotti F, Rossi C, Cicalini I, Di Santo S, Zucchelli M, Sacchetta P, Genovesi D, Pieragostino D. Serum lipidomic study reveals potential early biomarkers for predicting response to chemoradiation therapy in advanced rectal cancer: A pilot study. Adv Radiat Oncol. 2017 Jan 5;2(2):118-124. doi: 10.1016/j.adro.2016.12.005. eCollection 2017 Apr-Jun. Pieragostino D, D'Alessandro M, di Ioia M, Rossi C, Zucchelli M, Urbani A, Di Ilio C, Lugaresi A, Sacchetta P, Del Boccio P. An integrated metabolomics approach for the research of new cerebrospinal fluid biomarkers of multiple sclerosis. Mol Biosyst. 2015 Jun;11(6):1563-72. doi: 10.1039/c4mb00700j. Pieragostino D, Agnifili L, Fasanella V, D'Aguanno S, Mastropasqua R, Di Ilio C, Sacchetta P, Urbani A, Del Boccio P. Shotgun proteomics reveals specific modulated protein patterns in tears of patients with primary open angle glaucoma naïve to therapy. Mol Biosyst. 2013 Jun;9(6):1108-16. doi: 10.1039/c3mb25463a. Epub 2013 Apr 12. Pieragostino D, Bucci S, Agnifili L, Fasanella V, D'Aguanno S, Mastropasqua A, Ciancaglini M, Mastropasqua L, Di Ilio C, Sacchetta P, Urbani A, Del Boccio P. Differential protein expression in tears of patients with primary open angle and pseudoexfoliative glaucoma. Mol Biosyst. 2012 Apr;8(4):1017-28. doi: 10.1039/c1mb05357d. Epub 2011 Nov 29. Pieragostino, D. *; Del Boccio, P.; Di Ioia, M.; Pieroni, L.; Greco, V.; De Luca, G.; D'Aguanno, S.; Rossi, C.; Franciotta, D.; Centonze, D.; Sacchetta, P.; Di Ilio, C.; Lugaresi, A.; Urbani, A.,Oxidative modifications of cerebral transthyretin are associated with multiple sclerosis. Proteomics. 2013;13(6):1002-9. doi: 10.1002/pmic.201200395. Del Boccio P, Pieragostino D, Di Ioia M, Petrucci F, Lugaresi A, De Luca G, Gambi D, Onofrj M, Di Ilio C, Sacchetta P, Urbani A. Lipidomic investigations for the characterization of circulating serum lipids in multiple sclerosis. J Proteomics. 2011 Nov 18;74(12):2826-36. doi: 10.1016/j.jprot.2011.06.023. Epub 2011 Jul 4.READ MORE
Extracellular vesicles
More recently, important results have been obtained studying circulating EVs in biofluids and cell media. Our group described acid sphingomyelinase-enriched exosomes in the CSF of MS patients, a finding that helped to understand the molecular mechanisms that allow the increase of neurotoxic ceramides and the decrease of sphingomyelin species in MS. We have also shown that in MS the protein cargo of CSF EVs is very similar to that of tear EVs. Our group, for the first time, highlighted neuronal and microglial EVs in the tears of MS patients, a finding that points to the possible passage of EVs from the CNS to a peripheral compartment. The characterization of the EVs proteomics cargo is under investigation in our lab using many other biological matrices, such as blood, urine, cell media and saliva. Our goal is to apply this knowledge to the diagnosis and prognosis of diverse pathological conditions, including, but not limited to, cancer, multiple sclerosis and ophthalmic diseases, and to the assessment of pharmacological responses.
Pieragostino D, Lanuti P, Cicalini I, Cufaro MC, Ciccocioppo F, Ronci M, Simeone P, Onofrj M, van der Pol E, Fontana A, Marchisio M, Del Boccio P. Proteomics characterization of extracellular vesicles sorted by flow cytometry reveals a disease-specific molecular cross-talk from cerebrospinal fluid and tears in multiple sclerosis. J Proteomics. 2019 Jul 30;204:103403. doi: 10.1016/j.jprot.2019.103403. Cufaro MC, Pieragostino D, Lanuti P, Rossi C, Cicalini I, Federici L, De Laurenzi V, Del Boccio P. Extracellular Vesicles and Their Potential Use in Monitoring Cancer Progression and Therapy: The Contribution of Proteomics. J Oncol. 2019 Jun 9;2019:1639854. doi: 10.1155/2019/1639854. Rossi C, Cicalini I, Cufaro MC, Agnifili L, Mastropasqua L, Lanuti P, Marchisio M, De Laurenzi V, Del Boccio P, Pieragostino D. Multi-Omics Approach for Studying Tears in Treatment-Naïve Glaucoma Patients. Int J Mol Sci. 2019 Aug 18;20(16). pii: E4029. doi: 10.3390/ijms20164029. Pieragostino D, Cicalini I, Lanuti P, Ercolino E, di Ioia M, Zucchelli M, Zappacosta R, Miscia S, Marchisio M, Sacchetta P, Onofrj M, Del Boccio P. Enhanced release of acid sphingomyelinase-enriched exosomes generates a lipidomics signature in CSF of Multiple Sclerosis patients. Sci Rep. 2018 Feb 15;8(1):3071. doi: 10.1038/s41598-018-21497-5.READ MORE
Structural and functional studies on Acute Myeloid Leukemia (AML)
Our group has a long-lasting interest in the study of the molecular basis of acute myeloid leukemia (AML). Proteins of interest are investigated for their structure, function and interactions through the implementation of techniques ranging from x-ray crystallography, NMR, structural bioinformatics to fluorescence spectroscopy, circular dichroism and SPR. Knowledge gained in vitro is complemented by studies on cell lines and primary samples aimed at assessing the effect of investigational molecules. The main focus of research has been in the last years the main subtype of AML, i.e. that with mutations of the gene coding for nucleophosmin 1 (NPM1). We have characterized the multifaceted interaction properties of this protein by determining the structures of its complexes with both nucleic acids and other proteins. Additionally, we characterized the effect of AML-related mutations on the structure, subcellular localization and interactions of NPM1. Research is now focussed on the investigation of the effect of molecules that interfere with these associations. More recently we began to investigate the structure, function and interactome of an NPM1 binding partner , nucleostemin, a protein not expressed in differentiated cells but present in stem and cancer cells, including many AML subtypes.
De Cola A, Franceschini M, Di Matteo A, Colotti G, Celani R, Clemente E, Ippoliti R, Cimini AM, Dhez AC, Vallè B, Raineri F, Cascone I, Destouches D, De Laurenzi V, Courty J, Federici L (2018) N6L pseudopeptide interferes with nucleophosmin protein-protein interactions and sensitizes leukemic cells to chemotherapy. Cancer Lett 412: 272-282. Luchinat E, Chiarella S, Franceschini M, Di Matteo A, Brunori M, Banci L, Federici L (2018) Identification of a novel nucleophosmin-interaction motif in the tumor suppressor p14arf. FEBS J., 285(5):832-847 Di Matteo A, Franceschini M, Paiardini A, Grottesi A, Chiarella S, Rocchio S, Di Natale C, Marasco D, Vitagliano L, Travaglini-Allocatelli C, Federici L (2017) Structural investigation of nucleophosmin interaction with the tumor suppressor Fbw7γ. Oncogenesis. 6(9):e379. doi: 10.1038/oncsis.2017.78. Arcovito A, Chiarella S, Della Longa S, Di Matteo A, Lo Sterzo C, Scaglione GL, Federici L (2014) Synergic role of nucleophosmin three-helix bundle and a flanking unstructured tail in the interaction with G-quadruplex DNA. J. Biol. Chem. 289(31): 21230-21241. Chiarella S, De Cola A, Scaglione GL, Carletti E, Graziano V, Barcaroli D, Lo Sterzo C, Di Matteo A, DI Ilio C, Falini B, Arcovito A, De Laurenzi V, Federici L (2013) Nucleophosmin mutations alter its nucleolar localization by impairing G-quadruplex binding at ribosomal DNA. Nucleic Acids Res., 41(5): 3328-3339. Gallo A., Lo Sterzo C., Mori M., Di Matteo A., Bertini I., Banci L., Brunori M., Federici L. (2012) Structure of nucleophosmin DNA-binding domain and analysis of its complex with a G- quadruplex sequence from the c-MYC promoter. J. Biol. Chem., 287(32):26539-48 Federici L.*, Arcovito A., Scaglione G. L., Scaloni F., Lo Sterzo C., Di Matteo A., Falini B., Giardina B. and Brunori M. (2010) Nucleophosmin C-terminal leukaemia-associated domain interacts with G-rich quadruplex forming DNA. J. Biol. Chem., 285(48): 37138-37149.READ MORE
