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Your search keyword '"Spaggiari, G. M."' showing total 10 results
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10 results on '"Spaggiari, G. M."'

1. Multiparametric flow cytometry highlights B7-H3 as a novel diagnostic/therapeutic target in GD2neg/low neuroblastoma variants

Author

Dondero, A., Morini, M., Cangelosi, D., Mazzocco, K., Serra, M., Spaggiari, G. M., Rotta, G., Tondo, A., Locatelli, Franco, Castellano, A., Scuderi, F., Sementa, A. R., Eva, A., Conte, M., Garaventa, A., Bottino, C., Castriconi, R., Locatelli F. (ORCID:0000-0002-7976-3654), Dondero, A., Morini, M., Cangelosi, D., Mazzocco, K., Serra, M., Spaggiari, G. M., Rotta, G., Tondo, A., Locatelli, Franco, Castellano, A., Scuderi, F., Sementa, A. R., Eva, A., Conte, M., Garaventa, A., Bottino, C., Castriconi, R., and Locatelli F. (ORCID:0000-0002-7976-3654)

Abstract

Background High-risk neuroblastomas (HR-NBs) are rare, aggressive pediatric cancers characterized by resistance to therapy and relapse in more than 30% of cases, despite using an aggressive therapeutic protocol including targeting of GD2. The mechanisms responsible for therapy resistance are unclear and might include the presence of GD2neg/low NB variants and/or the expression of immune checkpoint ligands such as B7-H3. Method Here, we describe a multiparametric flow cytometry (MFC) combining the acquisition of 10 6 nucleated singlets, Syto16pos CD45neg CD56pos cells, and the analysis of GD2 and B7-H3 surface expression. 41 bone marrow (BM) aspirates from 25 patients with NB, at the onset or relapse, are analyzed, comparing results with cytomorphological analysis (CA) and/or immunohistochemistry (IHC). Spike in experiments assesses the sensitivity of MFC. Kaplan-Meier analysis on 498 primary NBs selects novel prognostic markers possibly integrating the MFC panel. Results No false positive are detected, and MFC shows high sensitivity (0.0005%). Optimized MFC identifies CD45negCD56pos NB cells in 11 out of 12 (91.6%) of BM indicated as infiltrated by CA, 7 of which coexpress high levels of GD2 and B7-H3. MFC detects CD45negCD56posGD2neg/low NB variants expressing high surface levels of B7-H3 in two patients with HR-NB (stage M) diagnosed at 53 and 139 months of age. One of them has a non-MYCN amplified tumor with unusual THpos PHOX2Bneg phenotype, which relapsed 141 months post-diagnosis with BM infiltration and a humerus lesion. All GD2neg/low NB variants are detected in patients at relapse. Kaplan-Meier analysis highlights an interesting dichotomous prognostic value of MML5, ULBPs, PVR, B7-H6, and CD47, ligands involved in NB recognition by the immune system. Conclusions Our study validates a sensitive MFC analysis providing information on GD2 and B7-H3 surface expression and allowing fast, specific and sensitive evaluation of BM tumor burden. With other routinely u

Published
2021

2. Dendritic cells efficiently cross-prime HLA class I-restricted cytolytic T lymphocytes when pulsed with both apoptotic and necrotic cells but not with soluble cell-derived lysates.

Author

Ferlazzo, G, Semino, C, Spaggiari, G M, Meta, M, Mingari, M C, and Melioli, G

Abstract

Dendritic cells (DC) have been shown to efficiently present antigen to CD8(+) cytolytic T cells (CTL) when pulsed with apoptotic cells as a source of cell-derived antigen. Such cross-priming could not be detected by the use of necrotic cells, while conflicting results have been reported for cell-derived soluble lysates. In this study, we reinvestigated this issue by using autologous Epstein-Barr virus-transformed lymphoblastoid cell lines (LCL) as a source of antigen to pulse monocyte-derived DC. Both autologous and HLA-mismatched allogeneic LCL have been used either in the form of apoptotic or of necrotic cells or of soluble cell lysates. At day 7, DC were co-cultured with the autologous CD8(+) lymphocyte fraction for an additional 9 days, in the presence of exogenous IL-2 (added after 48 h). At the end of the culture period, CD8(+) CTL efficiently lysed autologous LCL only when they had been co-cultured with DC pulsed with necrotic or apoptotic cells. That an efficient cross-priming of autologous CD8(+) cells could be induced by DC pulsed with apoptotic or necrotic LCL but not with cell lysates was further demonstrated in assays of IFN-gamma production in response to short-term re-stimulation of CD8(+) cells with LCL. In addition, LCL-specific CD8(+) cells could specifically lyse autologous DC that had been pulsed with LCL-derived antigens, further suggesting that DC presented exogenous antigens on HLA class I molecules.

Published
2000
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6. NK cell-mediated lysis of autologous antigen-presenting cells is triggered by the engagement of the phosphatidylinositol 3-kinase upon ligation of the natural cytotoxicity receptors NKp30 and NKp46.

Author

Spaggiari GM, Carosio R, Pende D, Marcenaro S, Rivera P, Zocchi MR, Moretta L, and Poggi A

Subjects
Antigens, Surface biosynthesis, Cells, Cultured, Histocompatibility Antigens Class I immunology, Humans, Killer Cells, Natural cytology, Natural Cytotoxicity Triggering Receptor 1, Natural Cytotoxicity Triggering Receptor 2, Natural Cytotoxicity Triggering Receptor 3, Phosphoinositide-3 Kinase Inhibitors, Receptors, Immunologic biosynthesis, Antigen-Presenting Cells immunology, Cytotoxicity, Immunologic immunology, Killer Cells, Natural immunology, Phosphatidylinositol 3-Kinases immunology, Receptors, Immunologic immunology
Abstract

Interleukin-2 (IL-2)-activated polyclonal or clonal NK cells lysed autologous antigen presenting cells (APC) through the engagement of the natural cytotoxicity receptors (NCR) NKp30 and NKp46. NK cell-mediated cytolysis of APC correlated with the surface density of these NCR. Indeed, NK cell clones bearing low amounts of NKp30 and NKp46 did not lyse autologous APC, whereas NK cell clones with bright expression of these NCR efficiently killed autologous APC. Upon masking of NKp30 or NKp46 by specific monoclonal antibodies a strong reduction (by 50%) of APC lysis could be detected and the complete inhibition was achieved by the simultaneous masking of these NCR. Interestingly, NK cell-mediated APC lysis was impaired by the phosphatidylinositol 3-kinase (PI-3 K) inhibitors LY294002 or wortmannin. Similarly, these drugs strongly reduced NK cell activation triggered by NKp30 or NKp46 in a re-directed killing assay as well as the activation of Akt/PKB, substrate of PI-3 K, induced by the engagement of these receptors. Altogether, these findings strongly suggest that NCR are responsible for the killing of autologous APC through the activation of PI-3 K.

Published
2001
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7. Engagement of CD33 surface molecules prevents the generation of dendritic cells from both monocytes and CD34+ myeloid precursors.

Author

Ferlazzo G, Spaggiari GM, Semino C, Melioli G, and Moretta L

Subjects
Antibodies, Monoclonal pharmacology, Cell Differentiation drug effects, Cells, Cultured, Dendritic Cells drug effects, Granulocyte-Macrophage Colony-Stimulating Factor pharmacology, Hematopoietic Stem Cells cytology, Hematopoietic Stem Cells drug effects, Hematopoietic Stem Cells immunology, Humans, Interleukin-4 pharmacology, Monocytes drug effects, Sialic Acid Binding Ig-like Lectin 3, Tumor Necrosis Factor-alpha pharmacology, Antigens, CD metabolism, Antigens, CD34 metabolism, Antigens, Differentiation, Myelomonocytic metabolism, Dendritic Cells cytology, Dendritic Cells immunology, Monocytes cytology, Monocytes immunology
Abstract

Although CD33 represents an important marker of myeloid cell differentiation, its function remains poorly defined. In view of its homology with p75/AIRM1, a recently identified surface molecule which exerts a potent inhibition on NK cell function, we re-evaluated the effect of CD33 engagement in defined myeloid cell functions. Addition of anti-CD33 mAb to cultures of CD14+ monocytes supplemented with granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-4 and TNF-alpha, prevented the generation of dendritic cells. In these cultured cells, engagement of CD33 resulted in an increased surface binding of annexin-V, followed by cell death. Mature dendritic cells were resistant to the CD33-mediated effect. Also in CD34+ precursors, cultured in the presence of flt3-ligand, c-Kit-ligand, GM-CSF, IL-4 and TNF-alpha, addition of anti-CD33 mAb prevented the recovery of mature dendritic cells. These data suggest a regulatory role of CD33 in the myeloid cell maturation and may offer a tool to interfere with the monocyte/macrophage cell function as well as with the development of dendritic cells.

Published
2000
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