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Patrizia BAROZZI
Personale tecnico amministrativo
Dipartimento di Scienze Mediche e Chirurgiche Materno-Infantili e dell'Adulto
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Pubblicazioni
- "Metodo per la diagnosi e/o il monitoraggio della mucormicosi" - "Method for the diagnosis of and/or monitoring mucormycosis"
[Brevetto]
Luppi, Mario; Potenza, Leonardo; Barozzi, Patrizia; Vallerini, Daniela; Forghieri, Fabio
abstract
A method is described for the diagnosis and/or monitoring of active or previous infection by Mucor which consists in the identification of Mucorales-specific T cells in samples from biological fluids taken from the patient and put into contact with a Mucor antigen. These specific immune responses can be detected by the execution of immunoenzymatic assays (ELISPOT, Quantiferon) or of immunocytofluorimetric assays [Cytokine Secretion Assay (CSA), Intracellular Cytokine Staining (ICS)] in vitro. In greater detail, the method in question provides for checking for the presence of specific IFN-γ producing T cells, of specific IL-10 producing T cells and/or specific IL- 4 producing T cells.
- Metodo per la diagnosi e/o il monitoraggio dell’ aspergillosi invasiva
[Brevetto]
Torelli, Giuseppe; Luppi, Mario; Barozzi, Patrizia; Potenza, Leonardo
abstract
La presente invenzione si riferisce ad un metodo per la diagnosi e/o ilmonitoraggio dell’infezione attiva o pregressa da Aspergillus fumigatus checomprende l’esecuzione di un saggio immunoenzimatico in vitro (ELISPOT) incui il campione da fluidi biologici prelevato dal paziente è messo in contatto conun antigene di Aspergillus fumigatus.
2023
- Characteristics and clinical behavior of acute myeloid leukemia harboring rare non-A/B/D nucleophosmin (NPM1) gene mutation subtypes: a single-center experience and review of the literature
[Articolo su rivista]
Mutti, M.; Cordella, S.; Parisotto, A.; Bettelli, F.; Morselli, M.; Cuoghi, A.; Bresciani, P.; Messerotti, A.; Gilioli, A.; Pioli, V.; Giusti, D.; Colaci, E.; Cassanelli, L.; Paolini, A.; Martinelli, S.; Maffei, R.; Riva, G.; Nasillo, V.; Sarti, M.; Trenti, T.; Comoli, P.; Tagliafico, E.; Manfredini, R.; Eccher, A.; Lagreca, I.; Barozzi, P.; Potenza, L.; Marasca, R.; Candoni, A.; Luppi, M.; Forghieri, F.
abstract
2023
- Improved efficacy of quizartinib in combination therapy with PI3K inhibition in primary FLT3-ITD AML cells
[Articolo su rivista]
Darici, Salihanur; Jørgensen, Heather G; Huang, Xu; Serafin, Valentina; Antolini, Ludovica; Barozzi, Patrizia; Luppi, Mario; Forghieri, Fabio; Marmiroli, Sandra; Zavatti, Manuela
abstract
: Acute myeloid leukemia is a heterogeneous hematopoietic malignancy, characterized by uncontrolled clonal proliferation of abnormal myeloid progenitor cells, with poor outcomes. The internal tandem duplication (ITD) mutation of the Fms-like receptor tyrosine kinase 3 (FLT3) (FLT3-ITD) represents the most common genetic alteration in AML, detected in approximately 30% of AML patients, and is associated with high leukemic burden and poor prognosis. Therefore, this kinase has been regarded as an attractive druggable target for the treatment of FLT3-ITD AML, and selective small molecule inhibitors, such as quizartinib, have been identified and trialled. However, clinical outcomes have been disappointing so far due to poor remission rates, also because of acquired resistance. A strategy to overcome resistance is to combine FLT3 inhibitors with other targeted therapies. In this study, we investigated the preclinical efficacy of the combination of quizartinib with the pan PI3K inhibitor BAY-806946 in FLT3-ITD cell lines and primary cells from AML patients. We show here that BAY-806946 enhanced quizartinib cytotoxicity and, most importantly, that this combination increases the ability of quizartinib to kill CD34+ CD38-leukemia stem cells, whilst sparing normal hematopoietic stem cells. Because constitutively active FLT3 receptor tyrosine kinase is known to boost aberrant PI3K signaling, the increased sensitivity of primary cells to the above combination can be the mechanistic results of the disruption of signaling by vertical inhibition.
2023
- Preclinical Validation of an Advanced Therapy Medicinal Product Based on Cytotoxic T Lymphocytes Specific for Mutated Nucleophosmin (NPM1mut) for the Treatment of NPM1mut-Acute Myeloid Leukemia
[Articolo su rivista]
De Cicco, Marica; Lagreca, Ivana; Basso, Sabrina; Barozzi, Patrizia; Muscianisi, Stella; Bianco, Alba; Riva, Giovanni; Di Vincenzo, Sara; Pulvirenti, Chiara; Sapuppo, Davide; Siciliano, Mariangela; Rosti, Vittorio; Candoni, Anna; Zecca, Marco; Forghieri, Fabio; Luppi, Mario; Comoli, Patrizia
abstract
2023
- Prognostic Relevance of Multi-Antigenic Myeloma-Specific T-Cell Assay in Patients with Monoclonal Gammopathies
[Articolo su rivista]
Lagreca, Ivana; Nasillo, Vincenzo; Barozzi, Patrizia; Castelli, Ilaria; Basso, Sabrina; Castellano, Sara; Paolini, Ambra; Maccaferri, Monica; Colaci, Elisabetta; Vallerini, Daniela; Natali, Patrizia; Debbia, Daria; Pirotti, Tommaso; Ottomano, Anna Maria; Maffei, Rossana; Bettelli, Francesca; Giusti, Davide; Messerotti, Andrea; Gilioli, Andrea; Pioli, Valeria; Leonardi, Giovanna; Forghieri, Fabio; Bresciani, Paola; Cuoghi, Angela; Morselli, Monica; Manfredini, Rossella; Longo, Giuseppe; Candoni, Anna; Marasca, Roberto; Potenza, Leonardo; Tagliafico, Enrico; Trenti, Tommaso; Comoli, Patrizia; Luppi, Mario; Riva, Giovanni
abstract
: Multiple Myeloma (MM) typically originates from underlying precursor conditions, known as Monoclonal Gammopathy of Undetermined Significance (MGUS) and Smoldering Multiple Myeloma (SMM). Validated risk factors, related to the main features of the clonal plasma cells, are employed in the current prognostic models to assess long-term probabilities of progression to MM. In addition, new prognostic immunologic parameters, measuring protective MM-specific T-cell responses, could help to identify patients with shorter time-to-progression. In this report, we described a novel Multi-antigenic Myeloma-specific (MaMs) T-cell assay, based on ELISpot technology, providing simultaneous evaluation of T-cell responses towards ten different MM-associated antigens. When performed during long-term follow-up (mean 28 months) of 33 patients with either MGUS or SMM, such deca-antigenic myeloma-specific immunoassay allowed to significantly distinguish between stable vs. progressive disease (p < 0.001), independently from the Mayo Clinic risk category. Here, we report the first clinical experience showing that a wide (multi-antigen), standardized (irrespective to patients' HLA), MM-specific T-cell assay may routinely be applied, as a promising prognostic tool, during the follow-up of MGUS/SMM patients. Larger studies are needed to improve the antigenic panel and further explore the prognostic value of MaMs test in the risk assessment of patients with monoclonal gammopathies.
2022
- Adverse outcome associated with daratumumab-based treatments in relapsed/refractory multiple myeloma patients with amplification of chromosome arm 1q21: a single-center retrospective experience
[Articolo su rivista]
Barbieri, Emiliano; Maccaferri, Monica; Leonardi, Giovanna; Giacobbi, Francesca; Corradini, Giorgia; Lagreca, Ivana; Barozzi, Patrizia; Potenza, Leonardo; Marasca, Roberto; Luppi, Mario
abstract
2020
- Acute myeloid leukemia in patients living with HIV infection: Several questions, fewer answers
[Articolo su rivista]
Forghieri, F.; Nasillo, V.; Bettelli, F.; Pioli, V.; Giusti, D.; Gilioli, A.; Mussini, C.; Tagliafico, E.; Trenti, T.; Cossarizza, A.; Maffei, R.; Barozzi, P.; Potenza, L.; Marasca, R.; Narni, F.; Luppi, M.
abstract
Both human immunodeficiency virus (HIV) infection and acute myeloid leukemia (AML) may be considered relatively uncommon disorders in the general population, but the precise incidence of AML in people living with HIV infection (PLWH) is uncertain. However, life expectancy of newly infected HIV-positive patients receiving anti-retroviral therapy (ART) is gradually increasing, rivaling that of age-matched HIV-negative individuals, so that the occurrence of AML is also expected to progressively increase. Even if HIV is not reported to be directly mutagenic, several indirect leukemogenic mechanisms, mainly based on bone marrow microenvironment disruption, have been proposed. Despite a well-controlled HIV infection under ART should no longer be considered per se a contraindication to intensive chemotherapeutic approaches, including allogeneic hematopoietic stem cell transplantation, in selected fit patients with AML, survival outcomes are still generally unsatisfactory. We discussed several controversial issues about pathogenesis and clinical management of AML in PLWH, but few evidence-based answers may currently be provided, due to the limited number of cases reported in the literature, mainly as case reports or small retrospective case series. Prospective multicenter clinical trials are warranted to more precisely investigate epidemiology and cytogenetic/molecular features of AML in PLWH, but also to standardize and further improve its therapeutic management.
2020
- BTK Inhibition Impairs the Innate Response Against Fungal Infection in Patients With Chronic Lymphocytic Leukemia
[Articolo su rivista]
Fiorcari, S.; Maffei, R.; Vallerini, D.; Scarfo, L.; Barozzi, P.; Maccaferri, M.; Potenza, L.; Ghia, P.; Luppi, M.; Marasca, R.
abstract
Infections represent a cause of morbidity and mortality in patients affected by chronic lymphocytic leukemia (CLL). Introduction of new drugs in CLL clinical practice has showed impressive efficacy, in particular those targeting BTK. Among the consistent clinical data, an increasing number of reports describing the occurrence of unexpected opportunistic fungal infections has been reported during treatment with ibrutinib in the first 6 months of treatment. The reason underlying manifestations of invasive fungal infections in patients treated with ibrutinib is still under investigation. Our study aimed to understand the impact of BTK inhibition on immune response to fungal infection mediated by macrophages and CD14+ monocytic population obtained from CLL patients. Exposure to ibrutinib and acalabrutinib reduced signaling pathways activated by Aspergillus fumigatus determining an exacerbation of an immunosuppressive signature, a reduction of phagocytosis and a significant deficit in the secretion of inflammatory cytokines either in macrophages and monocytes isolated from CLL patients and healthy donors. These effects lead to a failure in completely counteracting conidia germination. In addition we investigated the biological effects of ibrutinib on monocyte counterpart in patients who were undergoing therapy. A significant impairment in cytokine secretion and a deficit of phagocytosis in circulating monocytes were detected after 3 months of treatment. Thus, our results uncover modifications in the innate response in CLL patients induced by ibrutinib that may impair the immunological response to fungal infection. BTK inhibition affects a productive immune response of CLL-associated macrophages (NLC) during Aspergillus fumigatus infection. Reduction of TNF-α secretion and phagocytosis are detected in monocytes isolated from CLL patients during ibrutinib therapy.
2020
- Epidemiology and clinical outcomes of latent tuberculosis infection in adults affected with acute leukemia or aplastic anemia: a retrospective single-center study
[Articolo su rivista]
Bettelli, F.; Giusti, D.; Morselli, M.; Colaci, E.; Nasillo, V.; Pioli, V.; Gilioli, A.; Iotti, S.; Galassi, L.; Giubbolini, R.; Colasante, C.; Catellani, H.; Barozzi, P.; Lagreca, I.; Vallerini, D.; Maffei, R.; Franceschini, E.; Mussini, C.; Banchelli, F.; D'Amico, R.; Marasca, R.; Narni, F.; Potenza, L.; Comoli, P.; Luppi, M.; Forghieri, F.
abstract
2019
- Characterization and dynamics of specific T cells against nucleophosmin-1 (NPM1)-mutated peptides in patients with NPM1-mutated acute myeloid leukemia
[Articolo su rivista]
Forghieri, Fabio; Riva, Giovanni; Lagreca, Ivana; Barozzi, Patrizia; Vallerini, Daniela; Morselli, Monica; Paolini, Ambra; Bresciani, Paola; Colaci, Elisabetta; Maccaferri, Monica; Gilioli, Andrea; Nasillo, Vincenzo; Messerotti, Andrea; Pioli, Valeria; Arletti, Laura; Giusti, Davide; Bettelli, Francesca; Celli, Melania; Donatelli, Francesca; Corradini, Giorgia; Basso, Sabrina; Gurrado, Antonella; Cellini, Monica; Trenti, Tommaso; Marasca, Roberto; Narni, Franco; Martelli, Maria Paola; Falini, Brunangelo; Potenza, Leonardo; Luppi, Mario; Comoli, Patrizia
abstract
Nucleophosmin(NPM1)-mutated protein, a leukemia-specific antigen, represents an ideal target for AML immunotherapy. We investigated the dynamics of NPM1-mutated-specific T cells on PB and BM samples, collected from 31 adult NPM1-mutated AML patients throughout the disease course, and stimulated with mixtures of 18 short and long peptides (9-18mers), deriving from the complete C-terminal of the NPM1-mutated protein. Two 9-mer peptides, namely LAVEEVSLR and AVEEVSLRK (13.9-14.9), were identified as the most immunogenic epitopes. IFNγ-producing NPM1-mutated-specific T cells were observed by ELISPOT assay after stimulation with peptides 13.9-14.9 in 43/85 (50.6%) PB and 34/80 (42.5%) BM samples. An inverse correlation between MRD kinetics and anti-leukemic specific T cells was observed. Cytokine Secretion Assays allowed to predominantly and respectively identify Effector Memory and Central Memory T cells among IFNγ-producing and IL2-producing T cells. Moreover, NPM1-mutated-specific CTLs against primary leukemic blasts or PHA-blasts pulsed with different peptide pools could be expanded ex vivo from NPM1-mutated AML patients or primed in healthy donors. We describe the spontaneous appearance and persistence of NPM1-mutated-specific T cells, which may contribute to the maintenance of long-lasting remissions. Future studies are warranted to investigate the potential role of both autologous and allogeneic adoptive immunotherapy in NPM1-mutated AML patients.
2018
- Dual inhibition of PI3K/mTOR signaling in chemoresistant AMLprimary cells.
[Articolo su rivista]
Bertacchini, Jessika; Frasson, Chiara; Chiarini, Francesca; D'Avella, Daniele; Accordi, Benedetta; Anselmi, Laura; Barozzi, Patrizia; Foghieri, Fabio; Luppi, Mario; Martelli, Alberto M.; Basso, Giuseppe; Najmaldin, Saki; Khosravi, Abbas; Rahim, Fakher; Marmiroli, Sandra
abstract
A main cause of treatment failure for AML patients is resistance to chemotherapy. Survival of AML cells may depend on mechanisms that elude conventional drugs action and/or on the presence of leukemia initiating cells at diagnosis, and their persistence after therapy. MDR1 gene is an ATP-dependent drug efflux pump known to be a risk factor for the emergence of resistance, when combined to unstable cytogenetic profile of AML patients. In the present study, we analyzed the sensitivity to conventional chemotherapeutic drugs of 26 samples of primary blasts collected from AML patients at diagnosis. Detection of cell viability and apoptosis allowed to identify two group of samples, one resistant and one sensitive to in vitro treatment. The cells were then analyzed for the presence and the activity of P-glycoprotein. A comparative analysis showed that resistant samples exhibited a high level of MDR1 mRNA as well as of P-glycoprotein content and activity. Moreover, they also displayed high PI3K signaling. Therefore, we checked whether the association with signaling inhibitors might resensitize resistant samples to chemo-drugs. The combination showed a very potent cytotoxic effect, possibly through down modulation of MDR1, which was maintained also when primary blasts were co-cultured with human stromal cells. Remarkably, dual PI3K/mTOR inactivation was cytotoxic also to leukemia initiating cells. All together, our findings indicate that signaling activation profiling associated to gene expression can be very useful to stratify patients and improve therapy.
2018
- Effectiveness of originator (Neupogen) and biosimilar (Zarzio) filgrastim in autologous peripheral blood stem cell mobilization in adults with acute myeloid leukemia: a single-center retrospective study
[Articolo su rivista]
Nasillo, Vincenzo; Paolini, Ambra; Riva, Giovanni; Morselli, Monica; Potenza, Leonardo; Coluccio, Valeria; Maccaferri, Monica; Colaci, Elisabetta; Fantuzzi, Valeria; Messerotti, Andrea; Arletti, Laura; Pioli, Valeria; Lugli, Elisabetta; Gilioli, Andrea; Quadrelli, Chiara; Zucchini, Patrizia; Vallerini, Daniela; Lagreca, Ivana; Barozzi, Patrizia; Cuoghi, Angela; Bresciani, Paola; Marasca, Roberto; Mariano, Maria Teresa; Ceccherelli, Giovanni; Comoli, Patrizia; Campioli, Daniele; Trenti, Tommaso; Narni, Franco; Luppi, Mario; Forghieri, Fabio
abstract
n.d.
2017
- BCR-ABL-specific T-cell therapy in Ph+ ALL patients on tyrosine-kinase inhibitors
[Articolo su rivista]
Comoli, Patrizia; Basso, Sabrina; Riva, Giovanni; Barozzi, Patrizia; Guido, Ilaria; Gurrado, Antonella; Quartuccio, Giuseppe; Rubert, Laura; Lagreca, Ivana; Vallerini, Daniela; Forghieri, Fabio; Morselli, Monica; Bresciani, Paola; Cuoghi, Angela; Paolini, Ambra; Colaci, Elisabetta; Marasca, Roberto; Cuneo, Antonio; Iughetti, Lorenzo; Trenti, Tommaso; Narni, Franco; Foà, Robin; Zecca, Marco; Luppi, Mario; Potenza, Leonardo
abstract
Although the emergence of bone marrow (BM)-resident (p190)BCR-ABL-specific T lymphocytes has been correlated with hematologic and cytogenetic remissions in patients with Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph(+) ALL) undergoing maintenance tyrosine-kinase inhibitor treatment, little is known about the possibility of culturing these cells ex vivo and using them in T-cell therapy strategies. We investigated the feasibility of expanding/priming (p190)BCR-ABL-specific T cells in vitro by stimulation with dendritic cells pulsed with (p190)BCR-ABL peptides derived from the BCR-ABL junctional region and alternative splicing, and of adoptively administering them to patients with relapsed disease. We report on the feasibility of producing clinical-grade BCR-ABL-specific cytotoxic T lymphocytes (CTLs), endowed with antileukemia activity, from Ph(+) ALL patients and healthy donors. We treated 3 patients with Ph(+) ALL with autologous or allogeneic (p190)BCR-ABL-specific CTLs. No postinfusion toxicity was observed, except for a grade II skin graft-versus-host disease in the patient treated for hematologic relapse. All patients achieved a molecular or hematologic complete remission (CR) after T-cell therapy, upon emergence of (p190)BCR-ABL-specific T cells in the BM. Our results show that (p190)BCR-ABL-specific CTLs are capable of controlling treatment-refractory Ph(+) ALL in vivo, and support the development of adoptive immunotherapeutic approaches with BCR-ABL CTLs in Ph(+) ALL.
2017
- Detection of Fusarium-specific T cells in hematologic patients with invasive fusariosis
[Articolo su rivista]
Vallerini, Daniela; Forghieri, Fabio; Lagreca, Ivana; Riva, Giovanni; Barozzi, Patrizia; Quadrelli, Chiara; Morselli, Monica; Bresciani, Paola; Cuoghi, Angela; Coluccio, Valeria; Maccaferri, Monica; Paolini, Ambra; Colaci, Elisabetta; Marasca, Roberto; Narni, Franco; Cellini, Monica; Beauvais, Anne; Latgè, Jean Paul; Romani, Luigina; Iughetti, Lorenzo; Comoli, Patrizia; Campioli, Daniele; Trenti, Tommaso; Luppi, Mario; Potenza, Leonardo
abstract
N.A.
2017
- Multicenter Prospective Study for Laboratory Diagnosis of HHV8 Infection in Solid Organ Donors and Transplant Recipients and Evaluation of the Clinical Impact After Transplantation
[Articolo su rivista]
Chiereghin, Angela; Barozzi, Patrizia; Petrisli, Evangelia; Piccirilli, Giulia; Gabrielli, Liliana; Riva, Giovanni; Potenza, Leonardo; Cappelli, Gianni; de Ruvo, Nicola; Libri, Irene; Maggiore, Umberto; Morelli, Maria Cristina; Potena, Luciano; Todeschini, Paola; Gibertoni, Dino; Labanti, Manuel; Sangiorgi, Gabriela; la Manna, Gaetano; Pinna, Antonio Daniele; Luppi, Mario; Lazzarotto, Tiziana
abstract
BACKGROUND: We performed serological and molecular pretransplant screening in solid organ transplant (SOT) donors and recipients in north-central Italy and a surveillance program for human herpes virus 8 (HHV8) infection after transplant, aiming to establish an optimal management of HHV8 infection in SOT recipients. METHODS: For pretransplant HHV8 screening in both donors and recipients, 6 serological (4 indirect immunofluorescent assays (IFA) and 2 enzyme-linked immunosorbent assays (ELISA) - both HHV8 lytic and latent antigen-based) and 2 molecular assays were used. A reference standard to identify HHV8-positive patients was defined by at least 2 positive assays. All transplant patients at risk to develop HHV8-related disease underwent virological posttransplant monitoring by quantitative real-time PCR assay. RESULTS: HHV8 seroprevalence was 4% (10/249) in donors and 18% (93/517) in organ recipients. The best performance was obtained by 2 lytic antigen-based IFAs that showed almost perfect agreement to the reference standard (0.943 and 0.931 Cohen’s kappa). HHV8-DNA was detected in 6.8% and 2.9% of HHV8-seropositive donor samples by in-house nested PCR and quantitative real-time PCR assays, respectively. After transplant, 3 out of 12 (25%) HHV8-mismatch patients (seropositive donor/seronegative recipient) developed a primary infection, 1 of whom developed a lethal nonmalignant illness. Two out of 93 HHV8-seropositive recipients (2.1%) had viral replication in posttransplant period, 1 of whom developed Kaposi’s sarcoma. CONCLUSIONS: Serological assays, specifically lytic IFAs, were the best methodological approach to identify HHV8-infected SOT donors and recipients. A very low incidence (1.9%) of posttransplant HHV8-related disease was observed.
2017
- Successful Treatment of KSHV Inflammatory Cytokine Syndrome (KICS) after Kidney-Liver Transplant: Correlations with HHV8 miRNome and Specific T-Cell Response
[Articolo su rivista]
Mularoni, Alessandra; Gallo, Alessia; Riva, Giovanni; Barozzi, Patrizia; Miele, Monica; Cardinale, Giovanni; Vizzini, Giovanni; Volpes, Riccardo; Grossi, Paolo; Di Carlo, Daniele; Luca, Angelo; Trenti, Tommaso; Luppi, Mario; Conaldi, Pier Giulio
abstract
In post-transplant patients, HHV-8/KSHV infection is known to cause aggressive tumors, as well as severe non-neoplastic complications. These latter syndromes are driven by HHV-8/KSHV lytic reactivations and related hyper-inflammatory host responses, typically characterized by high viral loads, elevated levels of cytokines and other inflammation biomarkers, cytopenia, organ failure, high fever, and worsening conditions (with no evidence of B-cell neoplasias). These disorders are associated with a high mortality rate, often due to lack of prompt diagnosis, effective therapeutic approaches, and adequate follow up. Indeed, these features resemble most of those defining the so-called KSHV inflammatory cytokine syndrome (KICS), which was recently recognized in HIV-positive patients. Here we describe, for the first time, a case of KICS-like non-neoplastic recurrent complication occurring in an HIV-negative post-transplant patient, which was successfully treated by combination of anti-CD20 monoclonal therapy, antivirals, and modification of immunosuppressive regimen. In addition to clinical and laboratory findings collected during 3-year-long follow up, we also report novel experimental data on HHV-8-specific T-cell dynamics and circulating miRNA profile, showing correlations with clinical course and other laboratory markers (including viral load, CRP and cytokine levels), thus providing useful information about abnormal cellular and cytokine dynamics underlying of HHV-8-associated inflammatory disorders in post-transplant patients. This article is protected by copyright. All rights reserved.
2016
- All-trans retinoic acid (ATRA) in non-promyelocytic acute myeloid leukemia (AML): results of combination of ATRA with low-dose Ara-C in three elderly patients with NPM1-mutated AML unfit for intensive chemotherapy and review of the literature
[Articolo su rivista]
Forghieri, Fabio; Bigliardi, Sara; Quadrelli, Chiara; Morselli, Monica; Potenza, Leonardo; Paolini, Ambra; Colaci, Elisabetta; Barozzi, Patrizia; Zucchini, Patrizia; Riva, Giovanni; Vallerini, Daniela; Lagreca, Ivana; Marasca, Roberto; Narni, Franco; Venditti, Adriano; Martelli, Maria Paola; Falini, Brunangelo; Lo Coco, Francesco; Amadori, Sergio; Luppi, Mario
abstract
Based upon the clinical behavior of three patients, we suggest that the combination of low-dose Ara-C and all-trans retinoic acid may potentially be effective in some elderly patients, unfit for intensive chemotherapy, affected with NPM1-mutated acute myeloid leukemia without FLT3 mutations, warranting perspective clinical studies in these selected patients.
2016
- Chronic and recurrent benign lymphadenopathy without constitutional symptoms associated with human herpesvirus-6B reactivation
[Articolo su rivista]
Forghieri, Fabio; Luppi, Mario; Barozzi, Patrizia; Riva, Giovanni; Monica, Morselli; Bigliardi, Sara; Quadrelli, Chiara; Vallerini, Daniela; Maccaferri, Monica; Coluccio, Valeria; Paolini, Ambra; Colaci, Elisabetta; Goretta, Bonacorsi; Maiorana, Antonino; Sara, Tagliazucchi; Fabio, Rumpianesi; Mattioli, Francesco; Presutti, Livio; Gelmini, Roberta; Cermelli, Claudio; Giulio, Rossi; Patrizia, Comoli; Marasca, Roberto; Narni, Franco; Potenza, Leonardo
abstract
Chronic/recurrent behaviour may be encountered in some distinct atypical
or malignant lymphoproliferations, while recurrences are not generally
observed in reactive/benign lymphadenopathies. We retrospectively anal-
ysed a consecutive series of 486 human immunodeficiency virus-negative
adults, who underwent lymphadenectomy. Neoplastic and benign/reactive
histopathological pictures were documented in 299 (61 5%) and 187
(38 5%) cases, respectively. Of note, seven of the 111 (6 3%) patients with
benign lymphadenopathy without well-defined aetiology, showed chronic/
recurrent behaviour, without constitutional symptoms. Enlarged lymph
nodes were round in shape and hypoechoic, mimicking lymphoma. Reac-
tive follicular hyperplasia and paracortical expansion were observed.
Human herpesvirus (HHV)-6B positive staining in follicular dendritic cells
(FDCs) was documented in all seven patients. Serological, molecular and
immunological examinations suggested HHV-6B reactivation. Among the
remaining 104 cases with reactive lymphoid hyperplasia in the absence of
well-known aetiology and without recurrences, positivity for HHV-6B on
FDCs was found in three cases, whereas in seven further patients, a scanty
positivity was documented in rare, scattered cells in inter-follicular regions.
Immunohistochemistry for HHV-6A and HHV-6B was invariably negative
on 134 lymph nodes, with either benign pictures with known aetiology or
malignant lymphoproliferative disorders, tested as further controls. Future
studies are warranted to investigate a potential association between HHV-
6B reactivation and chronic/recurrent benign lymphadenopathy.
2016
- Circulating functional T cells specific to human herpes virus 6 (HHV6) antigens in individuals with chromosomally integrated HHV6
[Articolo su rivista]
Barozzi, Patrizia; Riva, Giovanni; Vallerini, Daniela; Quadrelli, Chiara; Lagreca, Ivana; Eccheli, Roberta; Forghieri, Fabio; Coluccio, Valeria; Maccaferri, Monica; Paolini, Ambra; Colaci, Elisabetta; Morselli, Monica; Marasca, Roberto; Narni, Franco; Potenza, Leonardo; Luppi, Mario; Comoli, Patrizia; Campioli, Daniele; Trenti, Tommaso
abstract
Circulating functional T cells specific to human herpes virus 6 (HHV6) antigens in individuals with chromosomally integrated HHV6
2016
- Mucorales-specific T cells in patients with hematologic malignancies
[Articolo su rivista]
Potenza, Leonardo; Vallerini, Daniela; Barozzi, Patrizia; Riva, Giovanni; Gilioli, Andrea; Forghieri, Fabio; Candoni, Anna; Cesaro, Simone; Quadrelli, Chiara; Maertens, Johan; Rossi, Giulio; Morselli, Monica; Codeluppi, Mauro; Mussini, Cristina; Colaci, Elisabetta; Messerotti, Andrea; Paolini, Ambra; Maccaferri, Monica; Fantuzzi, Valeria; DEL GIOVANE, Cinzia; Stefani, Alessandro; Morandi, Uliano; Maffei, Rossana; Marasca, Roberto; Narni, Franco; Fanin, Renato; Comoli, Patrizia; Romani, Luigina; Beauvais, Anne; Viale, Pier Luigi; Latgè, Jean Paul; Lewis, Russell E.; Luppi, Mario
abstract
Background: Invasive mucormycosis (IM) is an emerging life-threatening fungal infection. It is difficult to obtain a definite diagnosis and to initiate timely intervention. Mucorales-specific T cells occur during the course of IM and are involved in the clearance of the infection. We have evaluated the feasibility of detecting Mucorales-specific T cells in hematological patients at risk for IM, and have correlated the detection of such cells with the clinical conditions of the patients. Methods and Findings: By using an enzyme linked immunospot assay, the presence of Mucorales-specific T cells in peripheral blood (PB) samples has been investigated at three time points during highdose chemotherapy for hematologic malignancies. Mucorales-specific T cells producing interferon-γ, interleukin-10 and interleukin-4 were analysed in order to detect a correlation between the immune response and the clinical picture. Twenty-one (10.3%) of 204 patients, accounting for 32 (5.3%) of 598 PB samples, tested positive for Mucorales-specific T cells. Two groups could be identified. Group 1, including 15 patients without signs or symptoms of invasive fungal diseases (IFD), showed a predominance of Mucorales-specific T cells producing interferon-gamma. Group 2 included 6 patients with a clinical picture consistent with invasive fungal disease (IFD):2 cases of proven IM and 4 cases of possible IFD. The proven patients had significantly higher number of Mucorales-specific T cells producing interleukin-10 and interleukin-4 and higher rates of positive samples by using derived diagnostic cut-offs when compared with the 15 patients without IFD. Conclusions: Mucorales-specific T cells can be detected and monitored in patients with hematologic malignancies at risk for IM. Mucorales-specific T cells polarized to the production of T helper type 2 cytokines are associated with proven IM and may be evaluated as a surrogate diagnostic marker for IM.
2016
- PEGylated siRNA lipoplexes for silencing of BLIMP-1 in Primary Effusion Lymphoma: In vitro evidences of antitumoral activity
[Articolo su rivista]
Belletti, Daniela; Tosi, Giovanni; Forni, Flavio; Lagreca, Ivana; Barozzi, Patrizia; Pederzoli, Francesca; Vandelli, Maria Angela; Riva, Giovanni; Luppi, Mario; Ruozi, Barbara
abstract
Silencing of the B lymphocyte-induced maturation protein 1 (Blimp-1), a pivotal transcriptional regulator during terminal differentiation of B cells into plasma cells with siRNAs is under investigation as novel therapeutic approach in Primary Effusion Lymphoma (PEL), a HHV-8 related and aggressive B cell Lymphoma currently lacking of an efficacious therapeutic approach. The clinical application of small interfering RNA (siRNA) in cancer therapy is limited by the lack of an efficient systemic siRNA delivery system. In this study we aim to develop pegylated siRNA lipoplexes formed using the cationic lipid DOTAP and DSPE-PEG2000, capable to effectively stabilize anti-Blimp-1 siRNA and suitable for systemic administration. Two types of pegylated lipoplexes using a classic (C-PEG Lipoplexes) or a post-pegylation method (P-PEG-Lipoplexes) were formulated and compared in their physicochemical properties (size, zeta potential, morphology and structure) and efficiency on PEL cell lines. A stable siRNAs protection was obtained with post pegylation approach (2% molar of DSPE-PEG2000 with respect to lipid) resulting in structures with diameters of 300 nm and a complexation efficiency higher that 80% (0.08 nmol/10 nmol of lipid). In vitro studies on PEL cell lines suggested that empty liposomes were characterized by a low cell toxicity also after PEG modification (cell viability and cell density over 85% after treatment with 10 μM of lipid). We demonstrated that P-PEG-Lipoplexes were able to significantly reduce the levels of BLIMP-1 protein leading to reduction of viability (less that 15% after transfection with 100 nM of complexed siRNAs) and activation of apoptosis. In vitro efficiency encourages us to further test the in vivo potential of P-PEG-Lipoplexes in PEL therapy.
2016
- The bone marrow represents an enrichment site of specific T lymphocytes against filamentous fungi
[Articolo su rivista]
Vallerini, Daniela; Riva, Giovanni; Barozzi, Patrizia; Forghieri, Fabio; Lagreca, Ivana; Quadrelli, Chiara; Morselli, Monica; Bresciani, Paola; Cuoghi, Angela; Coluccio, Valeria; Maccaferri, Monica; Paolini, Ambra; Colaci, Elisabetta; Marasca, Roberto; Narni, Franco; Latgè, Jean Paul; Romani, Luigina; Comoli, Patrizia; Campioli, Daniele; Trenti, Tommaso; Luppi, Mario; Potenza, Leonardo
abstract
Bone marrow has already been described as an enrichment site for several antigen-specific T lymphocytes, but the presence of mould-specific T cells has never been investigated in the bone marrow. We have previously demonstrated that mould-specific T cells emerge in the peripheral blood of patients with invasive fungal infections (IFI) but tend to become undetectable after disease resolution. In seven patients with a history of IFI, we investigated the presence of mould-specific T cells secreting different cytokines in bone marrow and peripheral blood paired samples. The results showed that the frequencies of mould-specific T cells secreting the protective cytokine IFNI3 are significantly higher in bone marrow (BM) and are mainly represented by CD8+ T lymphocytes with effector phenotype. A putative disappearance of such protective BM responses after myeloablative therapy could contribute to the increased risk of IFI in hematologic patients.
2015
- Antineoplastic effects of liposomal siRNA treatment targeting BLIMP1/PRDM1 in primary effusion lymphoma
[Articolo su rivista]
Riva, Giovanni; Lagreca, Ivana; Mattiolo, Adriana; Belletti, Daniela; Lignitto, Laura; Barozzi, Patrizia; Ruozi, Barbara; Vallerini, Daniela; Quadrelli, Chiara; Corradini, Giorgia; Forghieri, Fabio; Marasca, Roberto; Narni, Franco; Tosi, Giovanni; Forni, Flavio; Vandelli, Maria Angela; Amadori, Alberto; Chieco Bianchi, Luigi; Potenza, Leonardo; Calabro', Maria Luisa; Luppi, Mario
abstract
RNA interference (RNAi) has been suggested to represent a promising therapeutic approach in different disease settings. Primary effusion lymphoma (PEL) is a plasmablastic lymphoma consistently expressing B lymphocyte-induced maturation protein 1 (Blimp-1), a pivotal transcriptional regulator during terminal differentiation of B cells into plasma cells. Here we report, for the first time, that transient knockdown of the BLIMP1 gene (also known as PR Domain Containing 1 with ZNF Domain, or PRDM1) using small interfering RNA (siRNA) delivered by liposomes, induced remarkable killing in PEL cell lines. Furthermore, in a murine model of PEL, significantly prolonged survival was achieved by intraperitoneal treatment with such anti-BLIMP1 lipoplexes, while no vector-induced toxicity was observed. This effective and safe RNAi strategy, based on liposomal siRNA targeting a master transcription factor of post-germinal center B cells, may indeed be a potential treatment against plasmablastic lymphoma
2015
- Epidemiology and clinical outcome of lower respiratory tract infections by respiratory syncytial virus or parainfluenza virus type 3 in adults receiving treatment for either acute leukemia or severe aplastic anemia: a retrospective single center study
[Articolo su rivista]
Bigliardi, Sara; Morselli, Monica; Potenza, Leonardo; Riva, Giovanni; Coluccio, Valeria; Maccaferri, Monica; Paolini, Ambra; Colaci, Elisabetta; Fantuzzi, Valeria; Soci, Francesco; Nasillo, Vincenzo; Messerotti, Andrea; Arletti, Laura; Pioli, Valeria; Lugli, Elisabetta; Gilioli, Andrea; Quadrelli, Chiara; Vallerini, Daniela; Barozzi, Patrizia; Lagreca, Ivana; Marasca, Roberto; Narni, Franco; Franceschini, Erica; Codeluppi, Mauro; Mussini, Cristina; Luppi, Mario; Forghieri, Fabio
abstract
Epidemiology and clinical outcome of lower respiratory tract infections by respiratory syncytial virus or parainfluenza virus type 3 in adults receiving treatment for either acute leukemia or severe aplastic anemia: a retrospective single center study
2015
- Immunologic evidence of a strong association between non-Hodgkin lymphoma and simian virus 40
[Articolo su rivista]
Tognon, Mauro; Luppi, Mario; Corallini, Alfredo; Taronna, Angelo; Barozzi, Patrizia; Rotondo, John Charles; Comar, Manola; Casali, Maria Vittoria; Bovenzi, Massimo; D'Agostino, Antonio; Vinante, Fabrizio; Rigo, Antonella; Ferrarini, Isacco; Barbanti Brodano, Giuseppe; Martini, Fernanda; Mazzoni, Elisa
abstract
Non-Hodgkin lymphoma (NHL), the most common cancer of the lymphatic system, is of unknown etiology. The identification of etiologic factors in the onset of NHL is a key event that could facilitate the prevention and cure of this malignancy. Simian virus 40 (SV40) has been considered an oncogenic agent in the onset/progression of NHL.
2015
- NPM1 mutations may reveal acute myeloid leukemia in cases otherwise morphologically diagnosed as myelodysplastic syndromes or myelodysplastic/myeloproliferative neoplasms
[Articolo su rivista]
Forghieri, Fabio; Paolini, Ambra; Morselli, Monica; Bigliardi, Sara; Bonacorsi, Goretta; Leonardi, Giovanna; Coluccio, Valeria; Maccaferri, Monica; Fantuzzi, Valeria; Faglioni, Laura; Colaci, Elisabetta; Soci, Francesco; Nasillo, Vincenzo; Messerotti, Andrea; Arletti, Laura; Pioli, Valeria; Zucchini, Patrizia; Quadrelli, Chiara; Corradini, Giorgia; Giacobbi, Francesca; Vallerini, Daniela; Riva, Giovanni; Barozzi, Patrizia; Lagreca, Ivana; Marasca, Roberto; Narni, Franco; Mecucci, Cristina; Ottaviani, Emanuela; Martinelli, Giovanni; Falini, Brunangelo; Luppi, Mario; Potenza, Leonardo
abstract
NPM1 mutations may reveal acute myeloid leukemia in cases otherwise morphologically diagnosed as myelodysplastic syndromes or myelodysplastic/myeloproliferative neoplasms.
2014
- Feedbacks and adaptive capabilities of the PI3K/Akt/mTOR axis in acute myeloid leukemia revealed by pathway selective inhibition and phosphoproteome analysis
[Articolo su rivista]
Bertacchini, Jessika; Guida, M; Accordi, B; Mediani, Laura; Martelli, A. M; Barozzi, Patrizia; Petricoin, E; Liotta, L; Milani, G; Giordan, M; Luppi, Mario; Forghieri, Fabio; DE POL, Anto; Cocco, L; Basso, G; Marmiroli, Sandra
abstract
Acute myeloid leukemia (AML) primary cells express high levels of phosphorylated Akt, a master regulator of cellular functions regarded as a promising drug target. By means of reverse phase protein arrays, we examined the response of 80 samples of primary cells from AML patients to selective inhibitors of the phosphatidylinositol 3 kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) axis. We confirm that >60% of the samples analyzed are characterized by high pathway phosphorylation. Unexpectedly, however, we show here that targeting Akt and mTOR with the specific inhibitors Akti 1/2 and Torin1, alone or in combination, result in paradoxical Akt phosphorylation and activation of downstream signaling in 70% of the samples. Indeed, we demonstrate that cropping Akt or mTOR activity can stabilize the Akt/mTOR downstream effectors Forkhead box O and insulin receptor substrate-1, which in turn potentiate signaling through upregulation of the expression/phosphorylation of selected growth factor receptor tyrosine kinases (RTKs). Activation of RTKs in turn reactivates PI3K and downstream signaling, thus overruling the action of the drugs. We finally demonstrate that dual inhibition of Akt and RTKs displays strong synergistic cytotoxic effects in AML cells and downmodulates Akt signaling to a much greater extent than either drug alone, and should therefore be explored in AML clinical setting.
2014
- Long-term molecular remission with persistence of BCR-ABL1-specific cytotoxic T cells following imatinib withdrawal in an elderly patient with Philadelphia-positive ALL
[Articolo su rivista]
Riva, Giovanni; Luppi, Mario; Lagreca, Ivana; Barozzi, Patrizia; Quadrelli, Chiara; Vallerini, Daniela; Zanetti, Eleonora; Basso, Sabrina; Forghieri, Fabio; Morselli, Monica; Maccaferri, Monica; Paolini, Ambra; Fantuzzi, Valeria; Messerotti, Andrea; Maffei, Rossana; Iacobucci, Ilaria; Martinelli, Giovanni; Marasca, Roberto; Narni, Franco; Comoli, Patrizia; Potenza, Leonardo
abstract
Long-term molecular remission with persistence of BCR-ABL1-specific cytotoxic T cells following imatinib withdrawal in an elderly patient with Philadelphia-positive ALL
2013
- Characterization of Specific Immune Responses to Different Aspergillus Antigens during the Course of Invasive Aspergillosis in Hematologic Patients
[Articolo su rivista]
Potenza, Leonardo; Vallerini, Daniela; Barozzi, Patrizia; Riva, Giovanni; Forghieri, Fabio; Beauvais, Anne; Beau, Remi; Candoni, Anna; Maertens, Johan; Rossi, Giulio; Morselli, Monica; Zanetti, Eleonora; Quadrelli, Chiara; Codeluppi, Mauro; Guaraldi, Giovanni; Pagano, Livio; Caira, Morena; DEL GIOVANE, Cinzia; Maccaferri, Monica; Stefani, Alessandro; Morandi, Uliano; Tazzioli, Giovanni; Girardis, Massimo; Delia, Mario; Specchia, Giorgina; Longo, Giuseppe; Marasca, Roberto; Narni, Franco; Merli, Francesco; Imovilli, Annalisa; Apolone, Giovanni; Carvalho, Agostinho; Comoli, Patrizia; Romani, Luigina; Latgè, Jean Paul; Luppi, Mario
abstract
Several studies in mouse model of invasive aspergillosis (IA) and in healthy donors have shown that different Aspergillus antigens may stimulate different adaptive immune responses. However, the occurrence of Aspergillus-specific T cells have not yet been reported in patients with the disease. In patients with IA, we have investigated during the infection: a) whether and how specific T-cell responses to different Aspergillus antigens occur and develop; b) which antigens elicit the highest frequencies of protective immune responses and, c) whether such protective T cells could be expanded ex-vivo. Forty hematologic patients have been studied, including 22 patients with IA and 18 controls. Specific T cells producing IL-10, IFN-γ, IL-4 and IL-17A have been characterized through enzyme linked immunospot and cytokine secretion assays on 88 peripheral blood (PB) samples, by using the following recombinant antigens: GEL1p, CRF1p, PEP1p, SOD1p, α1-3glucan, β1-3glucan, galactomannan. Specific T cells were expanded through short term culture. Aspergillus-specific T cells producing non-protective interleukin-10 (IL-10) and protective interferon-gamma (IFN-γ) have been detected to all the antigens only in IA patients. Lower numbers of specific T cells producing IL-4 and IL-17A have also been shown. Protective T cells targeted predominantly Aspergillus cell wall antigens, tended to increase during the IA course and to be associated with a better clinical outcome. Aspergillus-specific T cells could be successfully generated from the PB of 8 out of 8 patients with IA and included cytotoxic subsets able to lyse Aspergillus hyphae. Aspergillus specific T-cell responses contribute to the clearance of the pathogen in immunosuppressed patients with IA and Aspergillus cell wall antigens are those mainly targeted by protective immune responses. Cytotoxic specific T cells can be expanded from immunosuppressed patients even during the infection by using the above mentioned antigens. These findings may be exploited for immunotherapeutic purposes in patients with IA. © 2013 Potenza et al.
2013
- Human Herpesvirus 8 (HHV8) Infection and Related Diseases in Italian Transplant Cohorts.
[Articolo su rivista]
Riva, Giovanni; Barozzi, Patrizia; Quadrelli, Chiara; Vallerini, Daniela; Zanetti, Eleonora; Forghieri, Fabio; Chiereghin, A; Libri, I; Maggiore, U; Buzio, C; Lazzarotto, T; Narni, Franco; Luppi, Mario; Potenza, Leonardo
abstract
Human Herpesvirus 8 (HHV8) Infection and Related Diseases in Italian Transplant Cohorts
2013
- Trends in immune cell function assay and donor-specific HLA antibodies in kidney transplantation: A 3-year prospective study
[Articolo su rivista]
Libri, I; Gnappi, E; Zanelli, P; Reina, M; Giuliodori, S; Vaglio, A; Palmisano, A; Buzio, C; Riva, Giovanni; Barozzi, Patrizia; Luppi, Mario; Cravedi, P; Maggiore, U.
abstract
The immune cell function assay (ICFA) and de novo anti-donor-specific HLA antibodies (DSA) have been proposed as assays for immune monitoring in renal transplantation, but longitudinal studies examining the modification of both parameters over time and their relation with clinical events are lacking. We prospectively measured longitudinal changes in ICFA and DSA levels in 55 kidney transplant recipients over 3-year follow-up (534 visits) and analyzed their relation with the risk of developing acute rejections or infections. Seven patients (12.7%) developed biopsy-proven acute rejection, and 20 (36.4%) developed viral infections. At 3 years posttransplant, 28% of the patients had developed de novo DSA. ICFA levels peaked at 1-2 months posttransplant (p = 0.005) and leveled off thereafter. They were not associated with the risk of acute rejections, viral infections or development of de novo DSA. Instead, the incidence of de novo DSA was higher in patients who previously had viral infections (adjusted-odds ratio of de novo DSA associated with prior infections: 6.03 [95% CI, 1.64-22.06; p = 0.007]). Our prospective, longitudinal study does not support using ICFA to quantify the immune risk in kidney transplantation. Further studies are needed to confirm the relationship between viral infections and the subsequent development of de novo DSA.
2013
- siRNA-BASED THERAPEUTICS: DELIVERY AND TARGETING TO PEL TUMOR BY USING CATIONIC LIPOSOMES
[Abstract in Atti di Convegno]
Belletti, Daniela; Riva, Giovanni; Tosi, Giovanni; Lagreca, Ivana; Barozzi, Patrizia; Adriana, Mattiolo; Elena, Negri; Laura, Lignitto; Luigi Chieco, Bianchi; Forni, Flavio; Luppi, Mario; Vandelli, Maria Angela; Maria Luisa, Calabrò; Ruozi, Barbara
abstract
Aims of this research was to develop a “nanomedicine” approach based on siRNA delivery for the treatment of primary effusion lymphoma (PEL). The therapeutic use of antitumoral siRNA requires the development of specifically designed functional vectors, allowing improve¬ment of siRNA stability after systemic admin¬istration, and enabling targeted delivery directly into the neoplastic cells. In this context, liposomes, and particularly cationic liposomes, appears particu¬larly suitable to generate complexes with highly degradable siRNAs, as well as to specifically deliver siRNAs directly into the cytoplasm of the target tumor cells, where RNA interference processes take place. Generally, the electrostatic interaction between the positively charged lipids and the negatively charged nucleic acids leads to the formation of stable lipoplexes, protecting the cargo against nuclease attack and improving the cellular uptake and activity [1.
In this context, we are investigating innovative target strategies to improve the treatment of human herpesvirus 8 (HHV8)-associated primary effusion lymphoma (PEL). Primary effusion lymphoma (PEL) is an aggressive B cell non-Hodgkin’s lymphoma, affecting the serous cavities (such as the pleu¬ral, pericardial and abdominal cavities) and preferentially arising in immunocompromised or elderly patients, typically affected by several comorbidities and organ function impairments. PEL therapy has been revealed to be unsuccessful in the vast majority of patients, who are invariably characterized by a poor prognoses. Recently, small interfering RNAs (siRNAs), able to knock-down viral oncogenic proteins, were shown to induce efficient PEL cell apoptosis in vitro and PEL regressions in mice treated with intracavitary injection of lentiviral vectors expressing siRNA precursors[2.
Moving from our promising preliminary results in the field of nanotechnologies[3-4, we are developing different lipid-based nanocarriers (cationic and stealth-cationic liposomes), to deliver specific siRNAs to knock-down novel molecular targets (HHV8-encoded microRNAs, viral oncogenic proteins, or host transcription factors) with relevant functions in PEL pathogenesis [5. We are presently testing the delivery efficiency of these nanocarriers and the antineoplastic activity in vitro and in vivo using different PEL-derived cell lines and a previously established PEL mouse model[6.
We performed several preliminary technological experiments aimed at optimizing the operative condition to obtain the efficient liposomes/siRNAs complexes. Chemic-physical properties of both liposomes and lipoplexes were evaluated by exploiting microscopic, spectroscopic and gel electrophoresis techniques.
In vitro experiments demonstrated a high transfection efficiency of some of our carriers, which stably protected and efficaciously delivered siRNAs into PEL cells. Preliminary experiments using a mixture of siRNAs targeting a specific cellular gene showed a remarkable dose-dependent apoptosis, measured by annexin-V staining, in lipoplexes-transfected PEL cells. Moreover, the in vivo delivery of these therapeutic siRNAs significantly increased the survival time of treated mice compared with control treatment (log-rank test, lipoplexes vs empty liposomes, p=0.002), indicating that our lipoplexes exerted a significant antineoplastic activity. The empty carriers were not toxic in control mice and did not delay PEL development respect untraeted mice.
Our data indicate that our lipoplexes may therefore be considered as the basis for the development of useful short interfering RNA delivery vectors to treat PEL tumor. Moreover, we identified a target gene whose suppression exerts a relevant tumoricidal activity on PEL cells in vitro and in vivo, opening new perspectives for PEL treatment.
2012
- Atraumatic splenic rupture in patients with myelodysplastic syndromes: Report of a case occurred during treatment with 5-azacitidine and review of the literature.
[Articolo su rivista]
Forghieri, Fabio; Morselli, M; Leonardi, G; Potenza, Leonardo; Bonacorsi, G; Coluccio, V; Paolini, Ambra; Maccaferri, Monica; Colaci, Elisabetta; Fantuzzi, Valeria; Bigliardi, Sara; Zaldini, Piera; Riva, Giovanni; Barozzi, Patrizia; Leonardi, L; Rossi, A; Marasca, Roberto; Narni, Franco; Luppi, Mario
abstract
No abstract available
2012
- Chronic/relapsing lymphadenopathy associated with HHV-6B infection: a new benign clinico-pathologic entity occurring in immunocompetent individuals
[Abstract in Rivista]
Forghieri, Fabio; Potenza, Leonardo; Barozzi, Patrizia; Vallerini, Daniela; Riva, Giovanni; Zanetti, E; Quadrelli, C; Morselli, M; Leonardi, G; Maccaferri, M; Paolini, Ambra; Coluccio, Valeria; Colaci, Elisabetta; Pedrazzi, Letizia; Fantuzzi, Valeria; Bigliardi, Sara; Soci, Francesco; Bonacorsi, G; Zaldini, P; Rossi, G; Milani, M; Rivasi, Francesco; Gennari, W; Pecorari, M; Grottola, Antonella; Tagliazucchi, S; Rumpianesi, F; Mattioli, F; Presutti, Livio; Franzoni, Chiara; Gelmini, Roberta; Saviano, Massimo; Cermelli, Claudio; Marasca, Roberto; Narni, Franco; Luppi, Mario
abstract
Background. HHV-6 DNA sequences were disclosed in lymph node (LN) tis-
sues of several patients with lymphoid malignancies, but a direct major role of
HHV-6 in lymphoid malignant transformation has so far not been confirmed. In
contrast, active HHV-6 infection has been associated to either infectious
mononucleosis-like syndrome or acute lymphadenitis occurring in febrilepatients with systemic symptoms, or to Rosai-Dorfman disease in which viral
antigens have been detected by immunohistochimical (IHC) analyses in both
histiocytes and follicular dendritic cells (FDCs). Methods. We have retrospec-
tively analyzed clinical and pathological data of 365 adult patients, consecutive-
ly observed at our Institution over a period of 5 years (2006-2010), because of
enlarged superficial lymph nodes and subsequently undergoing lymphadenec-
tomy. In the benign/reactive cases in which well-recognized etiologies have
been excluded, an involvement of HHV-6 active infection or reactivation was
investigated by molecular and immunohistochemical examinations. Results.
Malignant disorders, namely malignant lymphoproliferative disorders or solid
cancer metastases, were found in 227 cases (62%), whereas in 138 cases
(38%) benign/reactive pictures were documented on lymph node examination.
Among these latter cases, a well-recognized etiology was demonstrated in 84
patients (61%), while in 54 cases (39%), a well-defined non-malignant
reactive/infectious cause could not be documented. Immunohistochemical
analyses resulted negative for both HHV-6A and HHV-6B in 38 of these latter
lymph nodes (70%). In 7 patients (13%), a scattered, scanty and aspecific pos-
itivity for HHV-6B late protein was documented in rare interfollicular plasma
cells and histiocytes. Surprisingly, in 9 patients (17%), immunohistochemical
analyses showed HHV-6B positive staining of FDCs, together with scattered
positivity of interfollicular cells. These 9 HIV-negative adult patients (median age
42 years, range 18-76 years), with either localized or generalized LAP, were
observed for a median follow-up of 38 months (range 28-166). Of note, six of
them presented with recurrent LAP (one to 3 recurrences), without evolving into
lymphoma. A common LN histological pattern at presentation showed florid fol-
licular hyperplasia with concurrent mild paracortical expansion. Three cases
also showed features consistent with PTGC. Constitutional symptoms were
absent in all patients. The IHC reactions for both HHV-6A and HHV-6B, per-
formed on further control cases, represented by 131 LN tissues from patients
with either benign LAP induced by other known etiologies or lymphoma, were
invariably negative. Serology was positive for both IgM and IgG with high avid-
ity suggesting viral reactivation/reinfection. However, the molecular analyses
failed to detect HHV-6 viremias in cell-free-serum samples of all the 9 patients
with positive HHV-6B IHC staining, while positivity for HHV-6B DNA was dis-
closed by PCR analyses in 7 out of the 7 LN tissues studied. Conclusions. We
show for the first time that local reactivation/infection of HHV-6B should be con-
sidered among the possible causes of chronic/relapsing benign LAP in immuno-
competent individuals. IHC is the method of choice for investigating the pres-
ence of HHV-6 infection in such cases. HHV-6B may indirectly modulate and
trigger the proliferation of lymphocytes, by locally affecting FDCs and LN
microenvironment. FDCs may indeed be involved in presenting HHV-6B anti-
gens to other immune cells, mainly cortical B lymphocytes.
2012
- How I treat HHV8/KSHV-related diseases in posttransplant patients.
[Articolo su rivista]
Riva, Giovanni; Luppi, Mario; Barozzi, Patrizia; Forghieri, Fabio; Potenza, Leonardo
abstract
Posttransplantation human herpesvirus-8 (HHV8)/Kaposi sarcoma herpesvirus (KSHV)
primary infection and/or reactivations are associated with uncommon and sometimes
fatal, neoplastic, and non-neoplastic diseases. HHV8-related clinical
manifestations notably range from Kaposi sarcoma (KS) to either primary effusion
lymphoma or multicentric Castleman disease B-cell malignancies, and from
polyclonal HHV8-positive plasmacytic lymphoproliferative disorders to bone marrow
failure and peripheral cytopenias, associated or not with hemophagocytic
syndromes, and to acute hepatitis syndromes. We reviewed the patient series
reported in the literature and summarized clinical management aspects, in terms
of diagnosis, follow-up, and treatment. We described typical clinical
presentations and histopathologic diagnostic features of these diseases, and we
discussed the role of HHV8-specific serologic, molecular, and immunologic assays,
particularly focusing on recent data from HHV8-specific T-cell monitoring in
posttransplantation KS patients. We finally discussed actual therapeutic options,
namely, the reduction or discontinuation of immunosuppressive therapy or the
switch from calcineurin inhibitors to mTOR inhibitors, as alternatives to
antineoplastic chemotherapy, along with the use of antiherpesvirus agents as
prophylactic or therapeutic measures, and treatment with rituximab in
posttrans-plantation multicentric Castleman disease patients and non-neoplastic
HHV8-associated syndromes.
2012
- LIPOPLEXES PER LA VEICOLAZIONE ED IL DIREZIONAMENTO DI siRNA AL PEL: VALUTAZIONE DELL’EFFICIENZA DI TRATTAMENTO CON SISTEMI LIPIDICI STABILIZZATI
[Relazione in Atti di Convegno]
Belletti, Daniela; Riva, Giovanni; Tosi, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela; Ruozi, Barbara
abstract
Per la stabilizzazione e la somministrazione di siRNAs, sono sempre più studiati vettori non virali, ed in particolare liposomi cationici, capaci di stabilizzare, per interazione elettrostatica, materiale genico generando strutture note come lipoplexes (Lpx). In questo contesto, sono stati formulati e testati Lpx attivi nel trattamento del PEL (Primary Effusion Lymphoma), un linfoma altamente aggressivo e con scarse possibilità terapeutiche. Inizialmente, liposomi allestiti con DOTAP sono stati impiegati per complessare 3 differenti siRNAs anti-BLIMP-1 (fattore trascrizionale fondamentale per mantenere lo stato neoplastico del PEL).
I Lpx ottenuti sono stati caratterizzati e ne è stata dimostrata l’elevata capacità trasfettiva sulla linea cellulare BCBL-1, modello di PEL. Per una somministrazione efficace, ed in previsione di studi in vivo, i complessi sono stati stabilizzati all’uptake macrofagico, attraverso modificazioni superficiali con PEG. Gli studi tecnologico-formulativi sono stati condotti sia partendo da liposomi pegilati che introducendo la pegilazione su complessi preformati. La complessazione tra liposomi cationici pegilati (preparati con DOTAP e DSPE-PEG) ed i siRNAs non è risultata efficace nello stabilizzare e trasferire il cargo alle cellule in coltura, presumibilmente a causa dell’ingombro e del tamponamento operato dalla copertura idrofila del liposoma, con conseguente limitata interazione tra il vettore ed il materiale genico. Incoraggianti risultatati sono invece derivati dall’applicazione della strategia “post complexation”, ovvero la pegilazione di Lpx cationici preformati (ottenuti tra liposomi DOTAP e siRNAs) mediante incubazione con micelle di DSPE-PEG. Tali “Post PEG-Lpx” hanno mostrato elevata capacità di stabilizzazione dell’attivo con conservata abilità transfettiva
2012
- Pathogenetic mechanisms of hepatitis C virus-induced B-cell lymphomagenesis.
[Articolo su rivista]
Forghieri, Fabio; Luppi, Mario; Barozzi, Patrizia; Maffei, Rossana; Potenza, Leonardo; Narni, Franco; Marasca, Roberto
abstract
Hepatitis C virus (HCV) infection is probably the most common chronic viral
infection and affects an estimated 180 million people worldwide, accounting for
3% of the global population. Although the liver is considered to be the primary
target, extrahepatic manifestations are well recognized among patients with
chronic HCV infection. Epidemiological studies have clearly demonstrated a
correlation between chronic HCV infection and occurrence of B-cell non-Hodgkin's
lymphomas (B-NHL). The clinical evidence that antiviral therapy has a significant
role in the treatment at least of some HCV-associated lymphoproliferative
disorders, especially indolent B-NHL, further supports the existence of an
etiopathogenetic link. However, the mechanisms exploited by HCV to induce B-cell
lymphoproliferation have so far not completely clarified. It is conceivable that
different biological mechanisms, namely, chronic antigen stimulation,
high-affinity interaction between HCV-E2 protein and its cellular receptors,
direct HCV infection of B-cells, and "hit and run" transforming events, may be
combined themselves and cooperate in a multifactorial model of HCV-associated
lymphomagenesis.
2012
- Successful generation of p190-BCR ABL-specific T-cell lines for prophylaxis/treatment of minimal residual disease in HSCT recipients with Ph+ acute lymphoblastic leukaemia
[Abstract in Rivista]
Basso, S.; Quartuccio, G.; Guido, I.; Quadrelli, C.; Gurrado, A.; Piantoni, L.; Zavras, N.; Cantoni, F.; Falcone, R.; Riva, Giovanni; Barozzi, Patrizia; Potenza, Leonardo; Maccario, R.; Zecca, M.; Luppi, Mario; Comoli, P.
abstract
Recent studies by our groups have demonstrated the presence of BM-homing, BCR-ABL specific cytotoxic T cells (CTL) in Philadelphia chromosome positive acute lymphoblastic leukemia (Ph+ALL) patients during sole Imatinib mesylate maintenance treatment, that inversely correlated with minimal residual disease (MRD). This observation supports the notion that anti-tumor T lymphocytes may effectively participate in the control of Ph+ leukemic proliferation, and represents the rationale for a BCR-ABL-targeted cell therapy approach to prophylactically treat leukemic relapse after HSCT in patients with Ph+ALL. The aim of this study was to evaluate the feasibility of expanding BCR-ABL specific CTL from HSCT donors, to be employed as specific DLI after HSCT for Ph+ALL. We conducted scale-up experiments to validate an in vitro culture method to expand BCR-ABL specific CTL from HSCT donors, by peripheral blood mononuclear cell (PBMC) stimulation with 9-20mer peptide pools derived from the p190 BCR-ABL fusion region. T-cell lines, that included a median 70% CD4+ and 29% CD8+ T lymphocytes, were successfully generated from 5/6 individuals. The T-cell lines showed specific INFg production in Elispot assays consistently higher (median 130 SFU/10e5 cells, range 0-198) than non-cultured donor PBMC (median 4 SFU/10e5 cells). In a standard 51chromium release assay, 5 of 6 T-cell lines presented specific cytotoxic activity against PHA blasts pulsed with BCR-ABL peptide mix (median lysis 30%, range 6-65), CD3-redirected activity against P815 cells (median lysis 38%, range 36-52) with minimal residual alloreactivity (median lysis 4%, range 0-15). Our data indicate that BCR-ABL-specific T-cell lines with limited alloreactivity may be expanded from HSCT donors after stimulation with BCR-ABL fusion region-derived peptides. Their efficacy in containment of MRD after allogeneic HSCT for Ph+ALL remains to be evaluated in clinical trials.
2011
- A case of JAK2 V617F-positive myelodysplastic/myeloproliferative neoplasm with unusual morphology, resembling acute promyelocytic leukemia-like disorder with a chronic course.
[Articolo su rivista]
Forghieri, Fabio; Morselli, M; Potenza, Leonardo; Maccaferri, Monica; Pedrazzi, Letizia; Coluccio, Valeria; Barozzi, Patrizia; Vallerini, Daniela; Riva, Giovanni; Zanetti, Eleonora; Quarelli, C; Bonacorsi, G; Artusi, Tullio; Zaldini, Piera; Zucchini, Patrizia; Marasca, Roberto; Narni, Franco; Falini, B; Torelli, Giuseppe; Luppi, Mario
abstract
An atypical case of myelodysplastic syndrome with morphologic aspect resempling acute promyelocytic leukemia, carring JA2 mutations.
2011
- BCR-ABL-specific cytotoxic T cells in the bone marrow of patients with Ph(+) acute lymphoblastic leukemia during second-generation tyrosine-kinase inhibitor therapy.
[Articolo su rivista]
Riva, Giovanni; Luppi, Mario; Quadrelli, Chiara; Barozzi, Patrizia; Basso, S; Vallerini, Daniela; Zanetti, E; Morselli, M; Forghieri, Fabio; Maccaferri, M; Paolini, Ambra; DEL GIOVANE, Cinzia; D'Amico, Roberto; Marasca, Roberto; Narni, Franco; Iacobucci, I; Martinelli, G; Baccarani, M; Comoli, P; Potenza, Leonardo
abstract
BCR-ABL-specific cytotoxic T cells in the bone marrow of patients with Ph+ acute lymphoblastic leukemia during second-generation tyrosine-kinase inhibitor therapy
2011
- Chronic eosinophilic leukaemia with ETV6-NTRK3 fusion transcript in an elderly patient affected with pancreatic carcinoma
[Articolo su rivista]
Forghieri, Fabio; Morselli, M; Potenza, Leonardo; Maccaferri, Monica; Pedrazzi, Letizia; Paolini, Ambra; Bonacorsi, G; Artusi, Tullio; Giacobbi, F; Corradini, G; Barozzi, Patrizia; Zucchini, Patrizia; Marasca, Roberto; Narni, Franco; Crescenzi, B; Mecucci, C; Falini, B; Torelli, Giuseppe; Luppi, Mario
abstract
Chronic eosinophilic leukaemia with ETV6-NTRK3 fusion transcript in an elderly patient affected with pancreatic carcinoma.A case report
2011
- Complessazione di SiRNA anti Blimp-1/PRDM con liposomi cationici: caratterizzazione chimico-fisica e validazione in vitro su cellule di PEL
[Relazione in Atti di Convegno]
Belletti, Daniela; Riva, Giovanni; Forni, Flavio; Barozzi, Patrizia; Luppi, Mario; Tosi, Giovanni; Vandelli, Maria Angela; Ruozi, Barbara
abstract
La tecnologia siRNAs (Small Interfering RNAs) è una innovativa strategia di regolazione genica post-trascrizionale potenzialmente applicabile in diversi campi della medicina ed in particolare in campo oncologico. [1] I siRNAs sono piccole sequenze di RNA a doppio filamento che a livello citoplasmatico subiscono un processo di attivazione mediato da un complesso sistema enzimatico denominato RISC (RNA induced silencing complexes) e divengono capaci di riconoscere e “silenziare” l’RNA target. In particolare le nostre ricerche sono indirizzate al linfoma effusivo delle cavità sierose (PEL). Si tratta di un raro tipo di linfoma di tipo non Hodgkin il cui agente eziologico è l’oncovirus HHV-8; nonostante i differenti approcci chemioterapici tentati, ad oggi non esiste un trattamento farmacologico efficace [2]. La disregolazione specifica mediata da siRNA del network di trascrizione della cellula malata potrebbe rappresentare una possibile alternativa nel trattamento della patologia. Considerando l’instabilità in vivo dei siRNAs associata alla loro scarsa capacità di penetrazione cellulare, il successo della strategia di silenziamento è strettamente dipendente dall’utilizzo di sistemi di delivery capaci di proteggere e trasportare selettivamente l’attivo nella cellula malata [3]. I liposomi, ed in particolare quelli cationici, sono sistemi di veicolazione e direzionamento innovativi, studiati e utilizzati da diversi anni, per la loro capacità di stabilizzare e trasferire materiale genico al bersaglio; la modificazione superficiale con molecole stabilizzanti (es PEG) e ligando selettive quali anticorpi monoclonali, peptidi ed aptameri garantisce la veicolazione e selettività del sistema [4,5].In studi preliminari, abbiamo dimostrato come sistemi liposomiali “stealth” modificati con anticorpo anti CD-138 specificamente riconosciuto da proteoglicani largamente espressi sulle cellule linfomatose, risultino altamente efficienti nella complessazione e stabilizzazione di materiale genico. Tali immunoliposomi sono stati testati quali carriers di un oligonucleotide modello (ODN-FITC) su cellule BCBL-1 (linea cellulare di PEL) evidenziando una buona capacità di trasferimento e di targeting valutata mediante citofluorimetria e microscopia confocale [6]. Tali evidenze sono risultate basilari per intraprendere nuovi studi di ottimizzazione di sistemi veicolanti siRNAs specifici nel silenziamento di fattori trascrizionali dominanti dello stadio plasmacellulare (knock-down di BLIMP-1/PRDM) in cellule PEL.In questa ricerca sono stati dapprima formulati e caratterizzati vettori liposomiali (Lp) utilizzando lipidi cationici (Dotap e DC-Chol) e neutri (Dope e Pc). Sono stati così selezionati i vettori che presentavano migliori caratteristiche in termini di dimensioni, carica superficiale e stabilità ed è stata valutata la loro capacità di complessare e stabilizzare siRNAs anti BLIMP-1. I complessi ottimizzati sono stati caratterizzati dal punto di vista chimico fisico mediante PCS e AFM ed è stata valutata l’efficienza di complessazione mediante elettroforesi su gel di agarosio. I dati hanno evidenziato come i liposomi allestiti con il lipide cationico Dotap risultino quelli maggiormente idonei alla formazione di lipoplexes idonei alla somministrazione: caratterizzati da strutture definite e riproducibili, di dimensioni prossime a 350nm che efficientemente proteggono i siRNAs, mostrando inoltre una buona stabilità dopo incubazione in siero. I lipoplexes allestiti con Dotap sono inoltre stati testati in vitro sulla linea cellulare BCBL-1, tali vettori sono risultati atossici in un ampio range di concentrazioni ed abili nel trasferire il materiale genico a livello citoplasmatico. In particolare è stato valutato l’effetto mediato dall’inibizione della proteina bersaglio mediante determinazione della proliferazione cellulare (analisi citomorfologica e conta cellulare) e test annessina/propidio per valutare
2011
- Differences in Kaposi sarcoma-associated herpesvirus-specific and herpesvirus-non-specific immune responses in classic Kaposi sarcoma cases and matched controls in Sicily.
[Articolo su rivista]
Amodio, E; Goedert, Jj; Barozzi, Patrizia; Riva, Giovanni; Firenze, A; Bonura, F; Viviano, E; Romano, N; Luppi, Mario
abstract
Kaposi sarcoma (KS) might develop because of incompetent immune responses, both non-specifically and specifically against the KS-associated herpesvirus (KSHV). Peripheral blood mononuclear cells from 15 classic (non-AIDS) KS cases, 13 KSHV seropositives (without KS) and 15 KSHV-seronegative controls were tested for interferon-γ T-cell (enzyme-linked immunospot [Elispot]) responses to KSHV-latency-associated nuclear antigen (LANA), KSHV-K8.1 and CMV/Epstein-Barr virus (EBV) peptide pools. The forearm and thigh of each participant was also tested for delayed-type hypersensitivity (DTH) against common recall antigens. Groups were compared with Fisher exact test and multinomial logistic regression to calculate odds ratios (OR) and 95% confidence intervals (CI). A KSHV Elispot response was detected in 10 (67%) classic KS cases, 11 (85%) KSHV seropositives (without KS) and two (13%) seronegative controls. All four cases with KSHV-LANA responses had current KS lesions, whereas five of six cases with KSHV-K8.1 responses had no lesions (P = 0.048). No case responded to both LANA and K8.1. Compared with the seronegative controls, the risk for classic KS was inversely related to DTH in the thigh (OR 0.71, 95% CI 0.55-0.94, P = 0.01), directly associated with DTH in the forearm (OR 1.35, 95% CI 1.02-1.80, P = 0.04) and tended to be increased fivefold per KSHV Elispot response (OR 5.13, 95% CI 0.86-30.77, P = 0.07). Compared with KSHV seropositives (without KS), the risk for classic KS was reduced fivefold (OR 0.20, CI 0.03-0.77, P = 0.04) per KSHV response. The CMV/EBV Elispot responses were irrelevant. Deficiency of both KSHV-specific and KSHV-non-specific immunity is associated with classic KS. This might clarify why Kaposi sarcoma responds to immune reconstitution.
2011
- Formulazione di siRNA anti blimp-1/PRDM in liposomi cationici: caratterizzazione chimico-fisica e validazione in vitro su cellule di LINFOMA EFFUSIVO PRIMARIO (pel)
[Abstract in Atti di Convegno]
Belletti, Daniela; Riva, Giovanni; Barozzi, Patrizia; Baraldi, Elisa; Veratti, Patrizia; Luppi, Mario; Tosi, Giovanni; Ruozi, Barbara
abstract
Formulazione di siRNA anti blimp-1/PRDM in liposomi cationici: caratterizzazione chimico-fisica e validazione in vitro su cellule di LINFOMA EFFUSIVO PRIMARIO (pel)
2011
- Identification and characterization of Aspergillus-specific immune responses to diagnose invasive aspergillosis in high risk patients: a multicenter study
[Abstract in Rivista]
Potenza, Leonardo; Vallerini, Daniela; Barozzi, Patrizia; Riva, Giovanni; Maertens, J; Candoni, A; Beauvais, A; Zanetti, Eleonora; Quadrelli, C; Morselli, M; Forghieri, Fabio; Maccaferri, M; Paolini, Ambra; Marasca, Roberto; DEL GIOVANE, Cinzia; D'Amico, Roberto; Ciceri, F; Comoli, P; Cesaro, S; Caira, M; Pagano, L; Romani, L; Narni, Franco; Latgè, Jp; Luppi, Mario
abstract
Background. The mortality of Invasive Aspergillosis (IA) still affects
from 27% to 55% of high risk hematologic patients. The reasons of
such a poor outcome also rely on several drawbacks limiting the di-
agnostic accuracy of non cultural based diagnostic methods
(NCBDM) and hampering the opportunities for an early intervention.
Studies in mice model of IA and in healthy subjects have shown that
Aspergillus-specific T-cells producing interferon-gamma (IFN-
gamma-T1) are protective, while Aspergillus-specific T-cells pro-
ducing interleukin-10 (IL-10-T2) are non-protective to IA. Aims. We
have investigated whether the identification of Aspergillus-specific
IFN-gamma-T1 and/or IL-10-T2 through an ex-vivo enzyme linked
immunospot (ELISPOT) assay may be effective in the diagnosis of IA
in high risk patients. Furthermore, in the proven IA patients, we have
functionally and phenotipically characterized such T cells through the
cytokine secretion assay (CSA). Methods. 180 patients (168 hemato-
logic and 12 solid organ transplant patients) have been enrolled. They
were classified, according the revised EORTC/MSG criteria, as fol-
lows: 18 proven, 35 probable, 17 possible IA cases and 110 controls.
The control patients were divided in two groups: group 1 included 86
(78.2%) patients with hystological and/or cultural verified infec-
tious/inflammatory/neoplastic diseases, but other than IA; group 2 in-
cluded 24 (21.8%) patients without clinical and/or microbiological
features of IA. ELISPOT has been performed, as described [Potenza et
al. Leukemia 2007; 21: 578-81], by using as antigens Aspergillus either
conidia or recombinant antigens, namely CRF1p, GEL1p, PEP1p,
SOD1p, α1-3 glucan, β1-3 glucan and galactomannan (GM). Results.
The patient and sample positivity rates were 94.4%/89.5% in proven,
45.7%/35.3% in probable, 35.3%/50% in possible IA cases and
1.8%/4.5% in the controls, respectively. The sensitivity and speci-
ficity of ELISPOT for the diagnosis of IA resulted 94.4% (95% CI,
73%-99%) and 98.2% (95% CI, 93%-99%), respectively. The PPV of
the test was 89.5% (95% CI, 67%-99%), the NPV was 99.1% (95%
CI, 94%-100%) and the efficiency was 97.6% (95% CI, 92.3%-
99.4%). The positive likelihood ratio (LR) resulted 51.89, the negative
LR was 0.06 (Table 1A,B). In proven IA patients, CSA demonstrated
that Aspergillus-specific IL-10-T2 were predominantly central
memory (CM) CD4+ T cells (median frequency 0.37%/0.22%), while
Aspergillus-specific IFN-gamma-T1 were either CD4+ or CD8+ cells
of either effector memory (EM) or CM phenotype (median frequen-
cies 0.24%/0.20%). Also lower frequencies of Aspergillus-specific ei-
ther CD4+ or CD8+ T cells producing IL-4 (0.11%/0.19%) of EM
phenotype, and EM CD8+ cells producing IL-17 (0.18%), were de-
tected. Moreover, although CRF1p, GEL1p, α1-3 glucan and SOD1p
resulted the antigens eliciting the highest number of Aspergillus-spe-
cific IFN-gamma-T1, only GEL1p and α1-3 glucan were those most
constantly targeted by protective immune responses along the entire
course of the IA. Conclusions. Our findings demonstrate the potential
of ELISPOT in the diagnosis of IA, suggesting that it may comple-
ment the other NCBDM, enabling a more consistent diagnosis of IA.
Furthermore, this study describes for the first time the Aspergillus-
specific immune responses in patients with proven IA, identifying
also the antigens predominantly targeted by protective IFN-gamma-
T1, with possible consequences in designing strategies of either
adoptive cell infusion or vaccine therapies.
2011
- KNOCK-DOWN DI BLIMP1/PRDM1 IN CELLULE PEL MEDIANTE siRNA; OTTIMIZZAZIONE DEL SISTEMA DI TRANSFEZIONE E STUDI IN VITRO
[Abstract in Atti di Convegno]
Belletti, Daniela; Riva, Giovanni; Tosi, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela; Ruozi, Barbara
abstract
Il trattamento del cancro mediante la chemioterapia tradizionale è spesso ostacolato dall’alta tossicità sistemica e dalla scarsa selettività dei principi attivi. La tecnologia siRNA (Small Interfering RNAs) basata sulla transfezione di ODN RNA antisenso, disegnati ad hoc per riconoscere e bloccare l’RNA messaggero target, rappresenta un’innovazione nelle strategie attuate in gene-silencing mostrando evidenti vantaggi anche in campo oncologico rispetto ai tradizionali chemioterapici. [1]Il linfoma effusivo delle cavità sierose (PEL), oggetto della ricerca, è un particolare tipo di linfoma associato invariabilmente all’infezione di HHV-8, con opzioni terapeutiche convenzionali limitate e non efficaci.[2] La disregolazione specifica mediata da siRNA del network di trascrizione della cellula malata rappresenta, ad oggi, una possibile alternativa nel trattamento della patologia. Considerando l’instabilità in vivo dei siRNAs associata alla loro scarsa capacità di penetrazione cellulare, il successo della strategia di silenziamento è strettamente dipendente dall’utilizzo di sistemi di delivery capaci di proteggere e trasportare selettivamente l’attivo nella cellula malata[3]. I liposomi, ed in particolare quelli cationici, sono sistemi di veicolazione e direzionamento innovativi, studiati e utilizzati da diversi anni, per la loro capacità di stabilizzare e trasferire materiale genico al bersaglio; la modificazione superficiale con molecole stabilizzanti (es PEG) e ligando selettive quali anticorpi monoclonali, peptidi ed aptameri garantisce la veicolazione e selettività del sistema [4,5].In studi preliminari, abbiamo dimostrato come sistemi liposomiali “stealth” modificati con anticorpo anti CD-138 specificamente riconosciuto da proteoglicani largamente espressi sulle cellule linfomatose, risultino altamente efficienti nella complessazione e stabilizzazione di materiale genico. Tali immunoliposomi sono stati testati quali carriers di un oligonucleotide modello (ODN-FITC) su cellule BCBL-1 (linea cellulare di PEL) evidenziando una buona capacità di trasferimento e di targeting valutata mediante citofluorimetria e microscopia confocale [6]. Tali evidenze sono risultate basilari per intraprendere nuovi studi di ottimizzazione di sistemi veicolanti siRNA specifici nel silenziamento di fattori trascrizionali dominanti dello stadio plasmacellulare (knock-down di BLIMP1/PRDM1) in cellule PEL. In questa ricerca sono stati formulati e caratterizzati vettori liposomiali (Lp) utilizzando lipidi neutri e cationici (Dotap e DC-Chol). Tali liposomi sono stati complessati con siRNAs e caratterizzati (elettroforesi su gel di agarosio, PCS ed AFM); i complessi ottenuti utilizzando il lipide cationico Dotap risultano caratterizzati da strutture definite e riproducibili, di dimensioni prossime a 350nm che efficientemente proteggono i siRNAs, mostrando inoltre una buona stabilità dopo incubazione in siero. I vettori sono stati testati in vitro su cellule BCBL-1; i dati preliminari evidenziano come i lipoplessi siano abili nel trasferire il materiale genico a livello citoplasmatico. L’inibizione della proteina bersaglio produce un effetto dose dipendente a concentrazioni superiori a 50nM evidente dopo 48/72 ore dal trattamento. In particolare è stata osservata un elevata induzione di necrosi /apoptosi mediante test annessina/propidio associata ad una significativa inibizione della proliferazione cellulare. E’ stata inoltre valutata la peghilazione come strategia per aumentare il tempo di permanenza in circolo oltre ad offrire il supporto per il legame con anticorpi specifici verso il PEL (anti.CD-138).[1] Oh YK, Park TG (2009). Adv. Drug Deliv Rev. 61: 850–862.[2] Carbone A, Gloghini A (2007). BJH review. 140: 13–24.[3] Whitehea KA , Langer R, Anderson DG (2009). Nature Review. 8:129-138.[4] Ruozi B, Belletti D, Tombesi A, Tosi G, Bondioli L, Forni F, Vandelli M A (2011). Int J Nanomed. 6:557–563.[5] Ruo
2011
- May the indirect effects of CIHHV-6 in transplant patients be exerted through the reactivation of the viral replicative machinery?
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Rossi, G; Riva, Giovanni; Vallerini, Daniela; Zanetti, Eleonora; Quadrelli, Chiara; Morselli, M; Forghieri, Fabio; Maccaferri, Monica; Paolini, Ambra; Marasca, Roberto; Narni, Franco; Luppi, Mario
abstract
Indirect effects of CIHHV-6 in transplant patients
2011
- Mucorles-specific T cells emerge in the course of invasive mucormucosis and may be used as a surrogate diagnostic marker in high-risk patients
[Articolo su rivista]
Potenza, Leonardo; Vallerini, Daniela; Barozzi, Patrizia; Riva, Giovanni; Forghieri, Fabio; Zanetti, Eleonora; Quadrelli, Chiara; Candoni, A; Maertens, J; Rossi, Giulio; Morselli, M; Codeluppi, M; Paolini, Ambra; Maccaferri, Monica; DEL GIOVANE, Cinzia; D'Amico, Roberto; Rumpianesi, F; Pecorari, M; Cavalleri, F; Marasca, Roberto; Narni, Franco; Luppi, Mario
abstract
Mucorales-specific T cells have been investigated in 28 hematologic patients during the course of their treatment. Three developed proven invasive mucormycosis (IM), 17 infections of known etiologies but other than IM, and 8 never showed fever upon the period of observation. The Mucorales-specific T cells may be detected only in patients with IM, at diagnosis and along the entire course of the IM, but neither before nor long time after the resolution of the infection. Such T cells produced predominantly interleukin-4, interferon-gamma (IFN-γ), interleukin-10, and to a lesser extent also interleukin-17, and belonged to either CD4+ or CD8+ subsets. The specific T cells producing IFN-γ were able to directly induce damage of Mucorales hyphae. None of the 25 patients without IM showed Mucorales-specific T cells. Specific T cells contribute to human immune responses against fungi of the order Mucorales and could be evaluated as a surrogate diagnostic marker of IM.
2011
- Novel polymeric/lipidic hybrid systems (PLHs) for effective Cidofovir delivery: preparation, characterization and comparative in vitro study with polymeric particles and liposomes
[Articolo su rivista]
Belletti, Daniela; Riva, Giovanni; Tosi, Giovanni; Forni, Flavio; Barozzi, Patrizia; Luppi, Mario; Vandelli, Maria Angela; Ruozi, Barbara
abstract
Cidofovir is an antiviral drug active as antitumoral agent a high doses against the Primary Effusion Lymphoma, a herpesvirus HHV8-associated B-cell lymphoma. A novel polymeric/lipidic hybrid system, consisting in a specific combination of biocompatible materials, capable to build a crossbred betweenpolymeric particles and liposomes were prepared and used to stabilize and deliver the drug, unsuccessfully formulated into several types of carriers. This innovative cidofovir-delivering system has structurally been characterized in comparison to multilamellar liposomes and polymeric particles, and then testedfor antitumoral efficacy against tumor cells (BCBL-1 cell line). The results demonstrated the improving of drug stability and encapsulation efficiency and suggested that polymeric/lipidic hybrid system could be promising to improve the antitumoral effect of cidofovir even at lower doses.
2011
- Tumor-targeted immunoliposomal nanosystems to deliver either Cidofovir or Antineoplastic SiRNA against Primary Effusion Lymphoma (PEL)
[Relazione in Atti di Convegno]
Riva, Giovanni; Belletti, Daniela; Ruozi, Barbara; Barozzi, Patrizia; Vallerini, Daniela; Quadrelli, Chiara; Zanetti, Eleonora; Morselli, M; Forghieri, Fabio; Marasca, Roberto; Narni, Franco; Tosi, Giovanni; Forni, Flavio; Vandelli, Maria Angela; Potenza, Leonardo; Luppi, Mario
abstract
Therapeutic applications of siRNA-mediated gene silencing appear to be highly dependent on the use of pharmaco-technologic carrier systems, able to protect siRNAs from rapid degradation upon administration, as well as to specifically deliver them to target cells. Actually, while siRNA-expressing viral vectors are burdened with safety concerns for their clinical use, the development of modified liposomal nanocarriers may represent a feasible option to harness the therapeutic potential of targetedantineoplastic siRNAs. Recently, we have successfully developed and characterized effective immunoliposomal nanosystems (ILNs) for targeted delivery of Cidofovir (an anti-herpesviral nucleotideanalogue, also showing antitumor activity) against PEL cell lines, demonstrating a significant improvement of the antineoplastic activity of the drug, especially at lower doses (less than 1nM). Thus, we tried to adapt such PEL-specific ILNs (PEGilated nanovescicles made of cationic/neutral lipids, engineered with anti-CD138 moAb on their surface) to efficiently encapsulate siRNAs and deliver them into PEL cell lines (highly expressing CD138 membrane protein). Our preliminary data showed thatsingle treatments with anti-PEL ILNs, delivering specific siRNAs against Blimp1 (Prdm1), which is a master transcription factor in PEL (a plasmablast/pre-plasmacell lymphoma, consistently Bcl-6 neg, Blimp1 pos), were able to induce a dose-dependent (50-200nM) inhibition of Blimp1 production (as assessed by Blimp1 mRNA and protein levels using RT-PCR and Western Blot, respectively), and this was strongly associated with enhanced cell death (more than 80%, using Annexin V/PI test). In particular, we observed a massive reduction of PEL viability (mean viable cells 8%, range 3-15%) as soon as 48-72 hours after treatment with 100nM anti-Blimp1 siRNAs. Interestingly, these data may resemble those described in multiple myeloma cell lines, after transduction with lentiviral vector constitutively expressing anti-Blimp1 shRNAs. Further studies on PEL murine models are now warranted to assess the efficacy and toxicity profile of in-vivo treatment with PEL-specific ILNs, loadedwith either Cidofovir or anti-Blimp1 siRNAs.
2011
- β-HHVs and HHV-8 in Lymphoproliferative Disorders.
[Articolo su rivista]
Quadrelli, Chiara; Barozzi, Patrizia; Riva, Giovanni; Vallerini, Daniela; E., Zanetti; Potenza, Leonardo; Forghieri, Fabio; Luppi, Mario
abstract
Similarly to Epstein-Barr virus (EBV), the human herpesvirus-8 (HHV-8) is a γ-herpesvirus, recently recognized to be associated with the occurrence of rare B cell lymphomas and atypical lymphoproliferations, especially in the human immunodeficiency virus (HIV) infected subjects. Moreover, the human herpesvirus-6 (HHV-6), a β-herpesvirus, has been shown to be implicated in some non-malignant lymph node proliferations, such as the Rosai Dorfman disease, and in a proportion of Hodgkin's lymphoma cases. HHV-6 has a wide cellular tropism and it might play a role in the pathogenesis of a wide variety of human diseases, but given its ubiquity, disease associations are difficult to prove and its role in hematological malignancies is still controversial. The involvement of another β-herpesvirus, the human cytomegalovirus (HCMV), has not yet been proven in human cancer, even though recent findings have suggested its potential role in the development of CD4(+) large granular lymphocyte (LGL) lymphocytosis. Here, we review the current knowledge on the pathogenetic role of HHV-8 and human β-herpesviruses in human lymphoproliferative disorders.
2010
- Cidofovir-loaded liposomes: an intro-study using BCBL-1 cell line as a model for primary effusion lymphoma
[Articolo su rivista]
Ruozi, Barbara; Riva, Giovanni; Belletti, Daniela; Tosi, Giovanni; Forni, Flavio; Mucci, Adele; Barozzi, Patrizia; Luppi, Mario; Vandelli, Maria Angela
abstract
Cidofovir (HPMPC) was recently reported to exert a valuable antineoplastic activity against primary effusion lymphoma (PEL), a B-cell neoplasm associated with Human Herpesvirus-8 (HHV-8) infection. In this study, we developed and characterized liposomes encapsulating HPMPC to increase drug efficacy reducing the administered dose and the related toxicity, which actually hamper its clinical therapeutic use in patients affected with PEL. The liposomes, obtained using different formulations of neutral and cationic lipids, were analyzed by microscopical (AFM) and spectroscopical (PCS and NMR) techniques. Using an in vitro model of PEL (BCBL-1 cell line), the carrier toxicity and the antineoplastic efficacy of liposomes were evaluated by flow cytometry applying apoptosis and cell death analysis. The in vitro study showed the applicability of the liposomes within a restricted range of lipidic concentrations according to the lipids used during the preparation. The moderate increases in the percentage of apoptotic/necrotic cells suggests that liposomal delivery allows the release of HPMPC into BCBL-1 cells enabling an unexpectedantineoplastic activity of this drug even at lower doses.
2010
- Common vascular endothelial growth factor variants and risk for posttransplant Kaposi sarcoma.
[Articolo su rivista]
Zanetti, E; Barozzi, Patrizia; Brown, Ee; Bosco, R; Vallerini, Daniela; Riva, Giovanni; Quadrelli, Chiara; Potenza, Leonardo; Forghieri, Fabio; Montagnani, G; D'Amico, Roberto; Del Giovane, Cinzia; Duraes, C; Whitby, D; Machado, Jc; Schulz, Tf; Torelli, G; Luppi, Mario
abstract
No abstract available
2010
- Development and characterization of immunoliposomes for Cidofovir and SiRNA delivery: a new strategy for the treatment of Primary Effusion Lymphoma
[Abstract in Atti di Convegno]
Belletti, Daniela; Ruozi, Barbara; Riva, Giovanni; Tosi, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela
abstract
Primary Effusion Lymphoma (PEL) is an aggressive and consistently lethal non-Hodgkin's B-cell lymphoma growing as lymphomatous effusions in serous body cavities and invariably associated with HHV-8 [1,2].The majority of the patients affected with PEL, either elderly HIV-negative or immunocompromised AIDS patients, are typically characterized by several age-related co-morbidities or opportunistic infectious diseases. Clinical efficacy of conventional anti-neoplastic chemotherapy is commonly hampered by the excessive grade of systemic toxicity and low drug levels in the tumor area [3].New therapeutic challenges may arise from the use of SiRNA technology, which is based on cellular transfection of antisense small RNAs, specifically designed to recognize the target mRNA and able to turn off the changed cellular mechanism of tumor cells, or by using antineoplastic drug (Cidofovir). Considering the high instability and the poor cellular uptake of both these actives, tumor-specific delivery by means of targeted nanocarriers is strongly required. These strategies represent an attractive approach to enhance intra-tumoral cytotoxic effects together with the reduction of “off-target” side effects, possibly offering a radical improvement in the treatment of such fragile oncologic patients. Among the colloidal carrier systems for drug delivery, liposomes have received considerable attention. They allow to protect the drug from rapid degradation, being particularly suitable to form complexes with highly-degradable ribonucleic acids.In this study, we formulated and characterized immunoliposomal formulation direct to PEL cell line (BCBL-1 cell line) using the cationic lipid DOTAP and a pegylated cholesterol funzionalized with a maleidoimide moiety capable to interact with anti CD-138 antibody. The formulation was characterized (size, zeta potential and morphology) in comparison with untargeted DOTAP liposomes and pegylated systems.These liposomal systems were used to transfect a model FITC-ODN into a model PEL cell line (BCBL-1). The studies on cellular binding and on the internalization of oligo by flow cytometry and confocal analysis confirmed the higher transfection efficiency of the immunoliposomes when compared with DOTAP and pegylated liposomes. This targeted formulation could be reasonably considered as optimal candidates for therapeutic siRNA delivery and more generally for gene encapsulation and delivery against the poor curable PEL tumor. Concerning cidofovir, it has been demonstrated that this antiviral drug is able to induce cell apoptosis in different tumor included PEL. Indeed, the high pro-apoptotic concentrations of cidofovir are never achievable in situ after full-dose systemic administration (5mg/Kg i.v.), and however, this systemic treatment can frequently cause severe nephrotoxicity [4]. We proposed liposomes encapsulating cidofovir by a modified reversed phase evaporation method (mREV) followed by extrusion. The characterization of samples suggested that cationic liposomes are more suitable for cidofovir stabilization, taking advantage of the charge interaction between the anionic drug and the cationic lipid moieties. Using the in vitro model of PEL (BCBL-1 cell line), the carrier toxicity and the antineoplastic efficacy of liposomes were evaluated by flow cytometry, applying apoptosis and cell death analysis. This in vitro study showed the applicability of the liposomes within a restricted range of lipidic concentrations, mainly depending on the lipids used during the preparation. The cidofovir transfection mediated by liposome composed of PC:DOTAP and PC:DC-CHOL caused a moderate increase in the percentage of apoptotic/necrotic PEL cells with respect to the controls (free drug and empty liposomes) suggesting that liposomal delivery allows the release of cidofovir into BCBL-1 cells enabling an unexpected antineoplastic activity of this drug even at lower doses.
2010
- Emergence of BCR-ABL-specific cytotoxic T cells in the bone marrow of patients with Ph+ acute lymphoblastic leukemia during long-term Imatinib mesylate treatment.
[Articolo su rivista]
Riva, Giovanni; Luppi, Mario; Barozzi, Patrizia; Quadrelli, Chiara; Basso, S; Vallerini, Daniela; Zanetti, Eleonora; Morselli, M; Forghieri, Fabio; Maccaferri, M; Volzone, F; DEL GIOVANE, Cinzia; D'Amico, Roberto; Locatelli, F; Torelli, Giuseppe; Comoli, P; Potenza, Leonardo
abstract
Imatinib mesylate (IM) has been demonstrated to be permissive for the emergence of T cells directed against chronic myeloid leukemia cells. Ten Philadelphia chromosome-positive acute lymphoblastic leukemia patients, receiving high dose IM maintenance, underwent a long-term immunological monitoring (range 2-65 months) of (p190)BCR-ABL-specific T cells in the bone marrow (BM) and peripheral blood (PB). (p190)BCR-ABL-specific T lymphocytes were detected in all patients, more frequently in BM than in PB samples (67% vs 25%, p<0.01), and resulted significantly associated with lower minimal residual disease values (p<0.001), while absent at leukemia relapse. Specific T cells were mainly effector memory CD8+ and CD4+ T cells, producing IFNgamma, TNFalpha and IL-2 (median % positive cells: 3.34, 3.04, 3.58, respectively). Cytotoxic subsets able to lyse BCR-ABL-positive leukemia blasts were also detectable. Whether these autologous (p190)BCR-ABL-specific T cells may be detectable under other tyrosine-kinase inhibitors, expanded ex vivo and exploited for immunotherapy remains to be addressed.
2010
- Formulation and characterization of new Polymeric/Lipidic Hybrid systems for cidofovir delivery
[Abstract in Atti di Convegno]
Belletti, Daniela; Riva, Giovanni; Tosi, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela; Ruozi, Barbara
abstract
Cidofovir is an antiviral drug with a remarkable antitumour activity in several animal models, associated or not with viral infections [1]. More recently, cidofovir has also been shown to be an effective treatment against Primary Effusion Lymphoma (PEL), a B cell non–Hodgkin lymphoma involving the serous cavities, invariably associated with Human HerpesVirus-8 (HHV8) and often with Epstein–Barr Virus (EBV) infection. A hindrance to the clinical applications of cidofovir is the high systemic toxicity, mainly the nephrotoxicity. Cidofovir encapsulation into specific micro- or nanocarriers, able to extend the release of the drug, may represent an effective strategy both to minimize the off-target organ exposure as well as to simultaneously increase the concentration of drug within the site of action. Unfortunately, the physical-chemical characteristics of the drug (low molecular weight, high solubility at different pH, unfavourable partition coefficient) limit the encapsulation into delivery systems [2,3]. Recently, we proposed the use of cationic liposomes to stably encapsulate cidofovir; unfortunately, the in vivo applicability of such cidofovir carriers is limited by the presence of cationic lipids, inducing a dose-related toxicity [4]. To overcome this problem, in this work we aimed to investigate a novel hybrid system, consisting in a specific combination of biocompatible materials, capable to build a crossbred between polymeric particles and liposomes. This innovative cidofovir-delivering systems (called PLHs, polymeric/lipidic hybrid systems) made of phosphatidilcholine (PC), cholesterol (CHOL) and polylactic acid (PLA) have been characterized (size, zeta potential, morphology, structure and thermal behaviour) in comparison to multilamellar liposomes and polymeric particles. Microscopical studies (atomic force and confocal microscopy), in agreement with the other characterizations, suggested that a rearrangement of the components has taken place to form a new matricial porous structure different both from liposomes and polymeric particles, with a wide dispersion of polymer in the lipidic bulk. This new crossbeald delivery systems was able to increased the encapsulation of cidofovir (encapsulation efficiency twice higher than liposomes and about 10 times higher than polymeric particles) and resulted atossic against PEL tumor cells (BCBL-1 cell line) revealing also a capability to better traslocate the drug into the cells causing and increased apoptosis respect to the free drug.
2010
- HHV-6 and atypical lymphoproliferative disorders: are only qualitative molecular examinations sufficient to support a pathogenetic role?
[Articolo su rivista]
Forghieri, Fabio; Potenza, Leonardo; Barozzi, Patrizia; Vallerini, Daniela; Riva, Giovanni; Zanetti, Eleonora; Quadrelli, Chiara; Torelli, Giuseppe; Luppi, Mario
abstract
n.d.
2010
- Immunoliposomal systems targeting the primary effusion lymphoma (PEL): in vitro study
[Articolo su rivista]
Ruozi, Barbara; Riva, Giovanni; Belletti, Daniela; Tosi, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela
abstract
Aims: To develop an appropriate liposomal formulation for gene delivery against Primary Effusion Lymphoma (PEL), a herpesvirus HHV8-associated B-cell lymphoma. Materials and methods: Cationic, cationic pegylated and cationic pegylated anti-CD138 liposomes (ILp) linking a monoclonal antibody expressed on PEL cells were prepared by thin layer evaporation method followed by extrusion technique. The formulations were mixed with a model oligonucleotide to form the lipoplexes tested on BCBL-1 cell (a PEL cell line). The transfection efficiency was evaluated by flow cytometry and confocal laser scanning microscopy analysis. Results: Based on antigen–antibody interaction, ILp mediated a specific gene delivery as shown by a significant increase in the transfection rate and a localized internalization of the oligo, in comparison with cationic liposomes and cationic pegylated liposomes.Conclusion: ILp could be proposed as effective carriers for oligo transfer in BCBL-1 cells. In vitro experimental results encourage to further test the in vivo therapeutic potentials of ILp for specific delivery of antitumoral agents.
2010
- Immunoliposomes for the delivery of SiRNA and chemioterapeutic agent to primary effusion lymphoma
[Abstract in Atti di Convegno]
Belletti, Daniela; Ruozi, Barbara; Riva, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela
abstract
In this study we have formulated and characterized immunoliposomes direct to PEL cells using the cationic lipid DOTAP and a pegylated cholesterol funzionalized with a maleidoimide moiety capable to stable interact with anti CD 138 antbody. The formulation was characterized in comparison with only pegylated systems and DOTAP liposomes. These liposomes were used to transfect a model of FITC-ODN into the target cells. The efficiency of these carriers was evaluated using flow cytometry and confocal analysis. The results confirm the higher transfection efficiency of the immunoliposomes if compared with conventional and pegylated liposomes.
2010
- Immunological and inflammatory features of Kaposi's sarcoma and other Kaposi's sarcoma-associated herpesvirus/human herpesvirus 8-associated neoplasias.
[Articolo su rivista]
Riva, Giovanni; Barozzi, Patrizia; Torelli, Giuseppe; Luppi, Mario
abstract
During the last 15 years, virologic and immunologic studies have provided a series of valuable clues on the modalities of gamma-herpesvirus-induced oncogenesis, which do not only consist of the direct subversion of intracellular signaling pathways, leading to a frank neoplastic molecular network in the infected cell, but also rely on viral manipulations of the cellular and cytokine microenvironment, especially in conditions of immunodeficiency in the host. At the virus-host interface, something iniquitous, strikingly favoring the aggressive expansion of human herpesvirus 8-infected lympho-endothelial clones, known as Kaposi's sarcoma, often occurs in different types of immunocompromised patients, able to establish a deleterious "pro-Kaposi's sarcoma" neo-angiogenic inflammatory network. However, these patients may control - or even resolve - the neoplastic burden as soon as an immunologic reassessment restores functional anti-Kaposi's sarcoma immune responses and reconstitutes a proper inflammatory environment. Indeed, the occurrence of iatrogenic Kaposi's sarcoma remissions, after the reduction or switch of immunosuppressive regimens, strongly suggests that the reset of immunologic constraints characterizing the Kaposi's sarcoma onco-pathogenic system may be sufficient to inhibit human herpesvirus 8-positive lympho-endothelial proliferations. Accordingly, immunologic reports all underline the pivotal protective role of anti-human herpesvirus 8 memory T-cells (harmonically, both CD8+ and CD4+ subsets), thus definitely implying a general requirement for an effective, antiviral immuno-inflammatory environment, based on correct and productive interactions between different compartments of dendritic, myeloid, and specific T-cells, in order to achieve and maintain optimal control on human herpesvirus 8-associated antigenic stimulations and Kaposi's sarcoma disease. In this review, we recapitulate some remarkable features about the outstanding immunologic issue raised by human herpesvirus 8-driven neoplastic outgrowths in immunodeficient patients, and in particular, we discuss the emerging view of Kaposi's sarcoma as an atypical neoplastic process, tightly dependent on immune system dynamics. It is conceivable that functional dissection of the specific immune responses, capable to cope with human herpesvirus 8, and further definitions of a global inflammatory profile with protective activity against Kaposi's sarcoma outbreaks, will eventually foster immunologic monitoring protocols during the follow-up of AIDS and posttransplant patients, either preventing or treating human herpesvirus 8-related tumors by multifunctional immunomodulation or prompt development of adoptive immunotherapeutic approaches.
2010
- Nanosistemi lipidici targettizzati per il direzionamento di siRNA con attività antineoplastica al linfoma primitivo delle cavità sierose (PEL)
[Abstract in Atti di Convegno]
Belletti, Daniela; Ruozi, Barbara; Riva, Giovanni; Tosi, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela
abstract
La tecnologia dei siRNA (Small Interfering RNAs) basata sulla transfezione di ODN RNA antisenso, disegnata ad hoc per riconoscere e bloccare l’RNA messaggero target, rappresenta l’innovazione nelle strategie attuate in gene-silencing con importanti prospettive per il trattamento di patologie complesse, come il linfoma primitivo delle cavità sierose (PEL), oggetto della ricerca.Il PEL è un particolare tipo di linfoma associato invariabilmente all’infezione di HHV-8, altamente aggressivo che presenta opzioni terapeutiche convenzionali limitate e non efficaci. La disregolazione specifica mediata da siRNA del network di trascrizione della cellula malata rappresenta, ad oggi, una possibile alternativa nel trattamento della patologia. I liposomi, ed in particolare quelli cationici, sono da diversi anni studiati per la loro capacità di stabilizzare materiale genico proteggendolo dalla degradazione in vivo ed offrendo inoltre la possibilità di coniugare molecole specifiche nel riconoscimento del target.In questa ricerca sono stati formulati e caratterizzati sistemi immunoliposomiali “stealth” e targettizzati con anticorpo CD-138 specificamente riconosciuto da proteoglicani largamente espressi sulle cellule linfomatose. Tali immunoliposomi sono stati testati in studi preliminari quali carriers di un oligonucleotide modello in una linea cellulare di PEL (BCBL-1); la capacità di trasferimento e di targeting è stata valutata mediante citofluorimetria e microscopia confocale. Successivamente sono stati allestiti immunoliposomi incorporanti siRNA per il silenziamento di fattori trascrizionali dominanti dello stadio plasmacellulare (knock-down di BLIMP1/PRDM1) e testati sulla stessa linea cellulare. I risultati evidenziano come i nostri sistemi siano altamente efficienti nella stabilizzazione del materiale genico promuovendo inoltre il trasferimento a livello citoplasmatico dove avviene il silenziamento della proteina bersaglio con conseguente apoptosi cellulare.
2010
- Organising pneumonia mimicking invasive fungal disease in patients with leukaemia.
[Articolo su rivista]
Forghieri, Fabio; Potenza, Leonardo; Morselli, M; Maccaferri, Monica; Pedrazzi, L; Barozzi, Patrizia; Vallerini, Daniela; Riva, Giovanni; Zanetti, Eleonora; Quadrelli, Chiara; Rossi, G; Rivasi, Francesco; Messino', M; Rumpianesi, F; Grottola, Antonella; Venturelli, C; Pecorari, M; Codeluppi, M; Torelli, Giuseppe; Luppi, Mario
abstract
Clinical charts from 63 consecutive highly immunocompromised haematologic patients presenting with pulmonary nodular lesions on CT scan, classified as either probable or possible invasive fungal disease (IFD) according to the revised EORTC/MSG classification, were retrospectively studied. Histopathological analysis of lung tissues, available for 23 patients, demonstrated proven IFD in 17 cases (14 invasive aspergillosis and 3 invasive zygomycosis), diffuse alveolar damage in one and organising pneumonia (OP) in five cases. In the OP cases, three of which have been defined as probable IFD according to EORTC/MSG classification, extensive immunohistochemical, molecular and immunological analyses for fungi were negative. Our case descriptions extend the notion that OP may be encountered as a distinct histopathological entity in pulmonary nodular lesions in patients with leukaemia with probable/possible IFD.
2010
- Splenic hyalohyphomycosis, molecularly and immunologically consistent with invasive aspergillosis, in a patient with acute lymphoblastic leukemia.
[Articolo su rivista]
Forghieri, Fabio; Rossi, G; Potenza, Leonardo; Morselli, M; Barozzi, Patrizia; Vallerini, Daniela; Messino, M; Rumpianesi, F; Pecorari, M; Torelli, Giuseppe; Luppi, Mario
abstract
n.d.
2010
- Translational challenges of human herpesvirus 6 chromosomal integration.
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Torelli, G; Luppi, Mario
abstract
No abstract available
2009
- Changes in T-Cell Responses Against Human Herpesvirus-8 Correlate with the Disease Course of Iatrogenic Kaposi's Sarcoma in a Patient with Undifferentiated Arthritis
[Articolo su rivista]
Barozzi, Patrizia; Potenza, Leonardo; Riva, Giovanni; Vallerini, Daniela; Quadrelli, Chiara; Bosco, Raffaella; Morselli, M; Forghieri, F; Volzone, F; Rossi, G; Ferri, Clodoveo; Bovini, C; Ciceri, F; Bordignon, C; Whitby, D; Schulz, Tf; Torelli, Giuseppe; Luppi, Mario
abstract
OBJECTIVES: To describe the first in-depth analysis of both the T-cell responses against human herpesvirus-8 (HHV-8) and the HHV-8 viral load in 1 patient who developed iatrogenic HHV-8-associated-Kaposi's sarcoma (KS) following immunosuppressive treatment for undifferentiated arthritis and to review the literature on iatrogenic KS (IKS). METHODS: T-cell responses against HHV-8 lytic and latent antigens were analyzed by ex vivo enzyme-linked immunospot (Elispot) and HHV-8 viral load was assessed by quantitative polymerase chain reaction, in sequential peripheral blood samples from a 55-year-old woman who developed skin/mucosal and visceral KS, while receiving treatment with cyclosporine, methotrexate, and methylprednisolone for undifferentiated arthritis. RESULTS: KS may result from HHV-8 infection in patients undergoing immunosuppressive treatment for rheumatic diseases and this is the first case of IKS occurring in undifferentiated arthritis. A role for immune surveillance in the pathogenesis of IKS is supported by the observation of disease regression following discontinuation of immunosuppressive therapy. In a 4-year follow-up, we showed that variations of the virus-specific immune responses but not of the viral load correlated well with the disease course, characterized by 2 remission and subsequent relapse phases, following changes of immunosuppressive therapy. CONCLUSIONS: We have provided evidence of a clear-cut correlation between changes in immunologic markers of HHV-8 infection and the disease course of this viral associated tumor, concomitant with variations of immunosuppressive treatment. Thus, ex vivo enzyme-linked immunospot for HHV-8-specific T-cell responses represents a new tool for the clinical management of rheumatic patients with IKS.
2009
- Diagnosis of aspergillosis: Role of proteomics
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Vallerini, Daniela; Zanetti, Eleonora; Torelli, Giuseppe; Luppi, Mario
abstract
The expansion of the antifungal armamentarium and the implementation of imaging techniques and new nonculture-based fungal diagnostics (NCBFDs) have improved the survival of patients with invasive aspergillosis (IA). However, mortality rates still remain high, possibly influenced by several pitfalls, affecting NCBFDs and reducing the window of opportunity for earlier treatment. A large body of in vitro and in vivo studies has demonstrated that several fungal proteic components are strongly immunogenic, and both the adaptive immunity and the innate branch are heavily involved in the recognition and clearance of fungal pathogens, resulting, on occasion, in a useful tool for the treatment of IA. By evaluating these studies, this review considers the possibility of exploiting either components of the innate or adaptive immunity to support the rapid and early diagnosis of IA. Copyright © 2009 by Current Medicine Group LLC.
2009
- Immunoliposomes for the delivery of SiRNA and chemioterapeutic agent to primary effusion lymphoma
[Abstract in Atti di Convegno]
Belletti, Daniela; Ruozi, Barbara; Riva, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela
abstract
Breve descrizione delle metodologie attuate per promuovere il rilascio di farmaci e materiale genico al tumore. In particolare è stato discusso l'approccio con liposomi e sistemi lipidici
2009
- Immunoliposomes to target the Primary Effusion Lymphoma (PEL): in vitro study
[Abstract in Atti di Convegno]
Belletti, Daniela; Ruozi, Barbara; Riva, Giovanni; Barozzi, Patrizia; Luppi, Mario; Forni, Flavio; Vandelli, Maria Angela
abstract
Preparazione e caratterizzazione di immunoliposomi per la veicolazione di materiale genico ai tumori (limfoma effusivo primario)
2009
- Indirect Antitumor Effects of Mammalian Target of Rapamycin Inhibitors Against Kaposi Sarcoma in Transplant Patients
[Articolo su rivista]
Barozzi, Patrizia; Riva, Giovanni; Vallerini, Daniela; Bosco, Raffaella; Quadrelli, Chiara; Zanetti, Eleonora; Potenza, Leonardo; Forghieri, Fabio; Torelli, Giuseppe; Luppi, Mario
abstract
n.d.
2009
- Interferon-alpha may restore sensitivity to tyrosine-kinase inhibitors in Philadelphia chromosome positive acute lymphoblastic leukaemia with F317L mutation
[Articolo su rivista]
Potenza, Leonardo; Volzone, F; Riva, Giovanni; Soverini, S; Martinelli, S; Iacobucci, I; Gnani, A; Barozzi, Patrizia; Forghieri, Fabio; Morselli, M; Zanetti, E; Maccaferri, Monica; Baccarani, M; Martinelli, G; Torelli, Giuseppe; Luppi, Mario
abstract
n.d.
2009
- Prevalence of Human Herpesvirus-6 Chromosomal Integration (CIHHV-6) in Italian Solid Organ and Allogeneic Stem Cell Transplant Patients
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Masetti, M; Pecorari, M; Bresciani, P; Gautheret Dejean, A; Riva, Giovanni; Vallerini, Daniela; Tagliazucchi, S; Codeluppi, M; DI BENEDETTO, Fabrizio; Gerunda, Giorgio Enrico; Narni, Franco; Torelli, Giuseppe; Luppi, Mario
abstract
The unique phenomenon of human herpesvirus-6 (HHV-6) chromosomal integration (CIHHV-6) may account for clinical drawbacks in transplant setting, being misinterpreted as active infection and leading to unnecessary and potentially harmful treatments. We have investigated the prevalence of CIHHV-6 in 205 consecutive solid organ (SO) and allogeneic stem cell transplant (alloSCT) Italian patients. Fifty-two (38.5%) of 135 solid organ transplant (SOT) and 16 (22.8%) of 70 alloSCT patients resulted positive for plasma HHV-6 DNA by real-time polymerase chain reaction. Seven SOT and three alloSCT patients presented HHV-6-related diseases, requiring antivirals. Two further patients (0.9%) were identified, presenting high HHV-6 loads. The quantification of HHV-6 on hair follicles disclosed the integrated state, allowing the discontinuation of antivirals. Before starting specific treatments, CIHHV-6 should be excluded in transplant patients with HHV-6 viremia by the comparison of HHV-6 loads on different fluids and tissues. Pretransplantation screening of donors and recipients may further prevent the misdiagnosis of CIHHV-6.
2008
- Acute appendicitis in adult neutropenic patients with hematologic malignancies
[Articolo su rivista]
Forghieri, Fabio; Luppi, Mario; Narni, Franco; Morselli, M.; Potenza, Leonardo; Bresciani, Paola; Volzone, Francesco; Rossi, Giulio; Rossi, Aldo; Trenti, Loris; Barozzi, Patrizia; Torelli, Giuseppe
abstract
n.d.
2008
- Assessment of aspergillus-specific T cells for diagnosis of invasive aspergillosis in a leukemic child with liver lesions mimicking hepatosplenic candidiasis
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Rossi, Giulio; Palazzi, G; Vallerini, Daniela; Riva, Giovanni; Cellini, M; Morselli, M; Volzone, Francesco; Venturelli, C; Quadrelli, Chiara; Di Pancrazio, L; Cano, Mc; Paolucci, Paolo; Torelli, Giuseppe; Luppi, Mario
abstract
A child with acute myeloid leukemia presented with multiple liver lesions mimicking hepatosplenic candidiasis during the neutropenic phase following the induction chemotherapy. All the available diagnostic tools showed repeatedly negative results, including galactomannan. An enzyme-linked immunospot (ELISPOT) assay showed a high number of Aspergillus-specific T cells producing interleukin-10 [TH2(IL-10)] and a low number of Aspergillus-specific T cells producing gamma interferon [TH1(IFN-γ)], revealing invasive aspergillosis (IA) before the confirmatory biopsy. A progressive skewing from the predominance of TH2(IL-10) to a predominance of TH1(IFN-γ) was observed close to the complete resolution of the infection and foreshadowed the outcome. The ELISPOT assay holds promise for diagnosing pediatric IA.
2008
- BK virus infection and neurologic dysfunctions in a patient with lymphoma treated with chemotherapy and rituximab
[Articolo su rivista]
Ferrari, A; Luppi, Mario; Marasca, Roberto; Potenza, Leonardo; Morselli, M; Volzone, F; Santachiara, R; Forghieri, Fabio; Barozzi, Patrizia; Torelli, Giuseppe
abstract
n.d.
2008
- Changes in the immune responses against human herpesvirus-8 in the disease course of posttransplant Kaposi sarcoma
[Articolo su rivista]
Barozzi, Patrizia; Bonini, C; Potenza, Leonardo; Masetti, Michele; Cappelli, Gianni; Gruarin, P; Whitby, D; Gerunda, Giorgio Enrico; Mondino, A; Riva, Giovanni; Vallerini, Daniela; Quadrelli, Chiara; Bosco, Raffaella; Ciceri, F; Bordignon, C; Schulz, Tf; Torelli, Giuseppe; Luppi, Mario
abstract
In nine patients with posttransplant Kaposi sarcoma (KS) T-cell responses to human herpesvirus (HHV)-8 latent and lytic antigens, as detected by enzyme-linked-immunospot (Elispot) assay, were absent at disease onset. Virus-specific T-cell responses were detected in six renal recipients at remission after a reduction of calcineurin inhibitors (CIs), and in two HHV-8 seropositive renal recipients without KS. In two liver recipients undergoing switch from CIs to sirolimus (SRL), normalization of the T-cell repertoire and recovery of both HHV-8-specific effector and memory T lymphocytes were associated with complete KS remission. In a renal recipient undergoing SRL conversion, the early recovery of HHV-8-specific effector but not of memory T lymphocytes, was associated only with partial remission. Neither rejection nor changes in graft function were observed after SRL conversion. HHV-8-specific T-cell responses are required to achieve posttransplant KS remission, and may be restored under SRL, while maintaining effective immunosuppression.
2008
- HHV-6A in syncytial giant-cell hepatitis
[Articolo su rivista]
Potenza, Leonardo; Luppi, Mario; Barozzi, Patrizia; Rossi, Giulio; Cocchi, Stefania; Codeluppi, Mauro; Pecorari, Monica; Masetti, Michele; DI BENEDETTO, Fabrizio; Gennari, William; Portolani, Marinella; Gerunda, Giorgio Enrico; Lazzarotto, Tiziana; Landini, Maria Paola; Schulz, Thomas F.; Torelli, Giuseppe; Guaraldi, Giovanni
abstract
Syncytial giant-cell hepatitis is a rare but severe form of hepatitis that is associated with autoimmune diseases, drug reactions, and viral infections. We used serologic, molecular, and immunohistochemical methods to search for an infectious cause in a case of syncytial giant-cell hepatitis that developed in a liver-transplant recipient who had latent infection with variant B of human herpesvirus 6 (HHV-6B) and who had received the organ from a donor with variant A latent infection (HHV-6A). At the onset of the disease, the detection of HHV-6A (but not HHV-6B) DNA in plasma, in affected liver tissue, and in single micromanipulated syncytial giant cells with the use of two different polymerase-chain-reaction (PCR) assays indicated the presence of active HHV-6A infection in the patient. Expression of the HHV-6A-specific early protein, p41/38, but not of the HHV-6B-specific late protein, p101, was demonstrated only in liver syncytial giant cells in the absence of other infectious pathogens. The same markers of HHV-6A active infection were documented in serial follow-up samples from the patient and disappeared only at the resolution of syncytial giant-cell hepatitis. Neither HHV-6B DNA nor late protein was identified in the same follow-up samples from the patient. Thus, HHV-6A may be a cause of syncytial giant-cell hepatitis.
2008
- Immunoliposome for the delivery of drugs and SiRNA for the treatment of cancer
[Abstract in Atti di Convegno]
Ruozi, Barbara; Riva, Giovanni; Barozzi, Patrizia; Tosi, Giovanni; Belletti, Daniela; Forni, Flavio; Luppi, Mario; Vandelli, Maria Angela
abstract
Immunoliposome for the delivery of drugs and SiRNA for the treatment of cancer
2008
- Parvoviruses in blood donors and transplant patients, Italy
[Articolo su rivista]
Vallerini, Daniela; Barozzi, Patrizia; Quadrelli, Chiara; Bosco, Raffaella; Potenza, Leonardo; Riva, Giovanni; Gregorini, G; Sandrini, S; Tironi, A; Montagnani, Giuliano; De Palma, M; Torelli, Giuseppe; Delwart, E; Luppi, Mario
abstract
n.d.
2008
- Veicolazione liposomiale del cidofovir nel trattamento del PEL (Primary Effusion Lynphoma)
[Abstract in Rivista]
Barozzi, Patrizia; Riva, Giovanni; Ruozi, Barbara; Tosi, Giovanni; Belletti, Daniela; Potenza, Leonardo; Quadrelli, Chiara; Vallerini, Daniela; Zanetti, Eleonora; M., Morselli; Forghieri, Fabio; Maccaferri, Monica; A., Paolini; F., Volzone; Vandelli, Maria Angela; Forni, Flavio; Torelli, Giuseppe; Luppi, Mario
abstract
Veicolazione liposomiale del cidofovir nel trattamento del PEL (Primary Effusion Lynphoma)
2007
- B cells and herpesviruses: a model of lymphoproliferation
[Articolo su rivista]
Barozzi, Patrizia; Potenza, Leonardo; Riva, Giovanni; Vallerini, Daniela; Quadrelli, Chiara; Bosco, Raffaella; Forghieri, Fabio; Torelli, Giuseppe; Luppi, Mario
abstract
Unlike alpha- and beta-herpesviruses, human gamma-herpesviruses, including the Epstein-Barr virus (EBV) and the human herpesvirus-8 (HHV-8), are B lymphotropic viruses. Primary infection with EBV, in otherwise healthy subjects, causes a benign lymphoproliferative syndrome, the mononucleosis syndrome. However, several epidemiologic and biologic studies have shown a pathogenetic role of EBV in the development of human B cell lymphomas, both in immunocompetent and in immunosuppressed individuals. HHV-8 is the necessary etiologic agent of a lymph vascular tumor, the Kaposi sarcoma, but it is also implicated in the pathogenesis of rare B cell lymphoproliferative disorders, mainly occurring in the setting of immunosuppression. The aim of this review is to provide an updated description of the different strategies used by these two herpesviruses to influence B cell fate decisions. Both EBV and HHV-8 have evolved specific mechanisms in order to: (1) interact with the B cell developmental machinery; (2) allow infected B cells to escape from the control of the immune system; (3) affect the B cell cycle checkpoints; (4) mimic and influence B cellular proliferation and differentiation pathways. Understanding the mechanisms of herpesvirus induced B cell lymphoproliferation will provide the basis for novel treatment approaches in patients with EBV and HHV-8 related lymphomas.
2007
- Diagnosis of invasive aspergillosis by tracking Aspergillus-specific T cells in hematologic patients with pulmonary infiltrates
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Vallerini, Daniela; Bosco, R; Quadrelli, Chiara; Morselli, M; Forghieri, Fabio; Volzone, F; Codeluppi, M; Rossi, G; Tazzioli, Giovanni; Venturelli, C; Torelli, Giuseppe; Luppi, Mario; Mediani, Laura
abstract
n.d.
2007
- Epstein-Barr virusassociated pneumonia in an adult patient with severe aplastic anaemia: resolutionafter the transient withdrawal of cyclosporine.
[Articolo su rivista]
Potenza, Leonardo; Barozzi, Patrizia; Codeluppi, M; Morselli, M; Forghieri, Fabio; Volzone, F; Riva, Giovanni; Pietrosemoli, P; Pecorari, M; Leonardi, G; Torelli, Giuseppe; Luppi, Mario
abstract
n.d.
2007
- Fatal hemophagocytic syndrome related to active human herpesvirus-8/Kaposi sarcoma-associated herpesvirus infection in human immunodeficiency virus-negative, non-transplant patients without related malignancies
[Articolo su rivista]
Re, A; Facchetti, F; Borlenghi, E; Cattaneo, C; Capucci, Ma; Ungari, M; Barozzi, Patrizia; Vallerini, Daniela; Potenza, Leonardo; Torelli, Giuseppe; Rossi, G; Luppi, Mario
abstract
Hemophagocytic syndrome (HS) may occur as a consequence of herpes viral infections. Human herpesvirus 8 (HHV-8)/Kaposi sarcoma-associated herpesvirus has so far been recognized as a trigger of HS only in immunosuppressed subjects or in patients with Kaposi sarcoma and/or HHV-8-related lymphoproliferative diseases. We report two Italian human immunodeficiency virus (HIV)-negative elderly men who developed an HS with a rapidly fatal course, following treatment with corticosteroids for autoimmune hemolytic anemia. An overwhelming active infection with HHV-8 was unequivocally documented by molecular and immunohistochemical methods, in the absence of HHV-8-related tumors. The occurrence of HHV-8-associated HS, although rare, may be considered, even out of the HIV or the transplantation settings, at least in areas endemic for HHV-8 infection.
2006
- Human herpesvirus 6 latency characterized by high viral load: Chromosomal integration in many, but not all, cells
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Bosco, Raffaella; Vallerini, Daniela; Potenza, Leonardo; Forghieri, Fabio; Torelli, Giuseppe
abstract
n.d.
2006
- KSHV/HHV-8 infection of tubular epithelial cells in transplantation kidney
[Articolo su rivista]
Barozzi, Patrizia; R., Bosco; Vallerini, Daniela; Potenza, Leonardo; Torelli, Giuseppe; Luppi, Mario; F., Facchetti; Guaraldi, Giovanni; T. F., Schulz
abstract
n.d.
2006
- Kaposi's sarcoma triggered by endogenous HHV-8 reactivation after non-myeloablative allogeneic haematopoietic transplantation
[Articolo su rivista]
Bruno, B; Sorasio, R; Barozzi, Patrizia; Vieira, J; Omede, P; Giaretta, F; Rotta, M; Giaccone, L; Massaia, M; Luppi, Mario; Boccadoro, M.
abstract
Human herpesvirus 8 (HHV-8) is causally associated with Kaposi's sarcoma (KS). KS is most frequently observed in HIV patients and in solid organ transplant recipients. The role of HHV-8 in allogeneic haematopoietic cell transplantation (HCT) remains to be determined. Here we describe a case in which KS concomitantly occurred with CMV reactivation after a non-myeloablative allogeneic HCT and presented with skin lesions, but not visceral involvement. Skin biopsy confirmed the diagnosis and ruled out graft versus host disease or disease recurrence. Molecular findings indicated viral reactivation of the recipient's primary infection. Tumour lesions completely receded when immunosuppression was tapered. Prevalence studies in donors and recipients are needed to determine the clinical impact of HHV-8 in HCT.
2005
- Suppressive effect of human herpesvirus-8/Kaposi sarcoma-associated herpesvirus on in vitro colony formation of hematopoietic progenitor cells
[Articolo su rivista]
Luppi, Mario; Trovato, R; Barozzi, Patrizia; Gibellini, F; Potenza, Leonardo; Riva, Giovanni; Torelli, Giuseppe
abstract
n.d.
2005
- Treatment of herpesvirus associated primary effusion lymphoma with intracavity cidofovir
[Articolo su rivista]
Luppi, Mario; Trovato, R; Barozzi, Patrizia; Vallisa, D; Rossi, Giorgio; Re, A; Ravazzini, L; Potenza, Leonardo; Riva, Giovanni; Morselli, M; Longo, G; Cavanna, L; Roncaglia, R; Torelli, Giuseppe
abstract
n.d.
2004
- Is it now the time to update treatment protocols for lymphomas with new anti-virus systems?
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Potenza, Leonardo; Riva, Giovanni; Morselli, M; Torelli, Giuseppe
abstract
n.d.
2003
- Erratum: Post-transplant Kaposi sarcoma originates from the seeding of donor-derived progenitors (Nature Medicine (2003) 9 (554-561))
[Articolo su rivista]
Barozzi, P.; Luppi, M.; Facchetti, F.; Mecucci, C.; Alu, M.; Sarid, R.; Rasini, V.; Ravazzini, L.; Rossi, E.; Festa, S.; Crescenzi, B.; Wolf, D. G.; Schulz, T. F.; Torelli, G.
abstract
2003
- Human herpesvirus 8-associated diseases in solid-organ transplantation: importance of viral transmission from the donor
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Guaraldi, Giovanni; L., Ravazzini; Rasini, Valeria; C., Spano; Riva, Giovanni; Vallerini, Daniela; Ad, Pinna; Torelli, Giuseppe
abstract
No abstract available
2003
- KSHV reactivation in post-transplant Kaposi sarcoma - Reply
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Facchetti, F; Schulz, Tf; Torelli, Giuseppe
abstract
No Abstract available
2003
- Mysticism to medicine
[Articolo su rivista]
Barozzi, Patrizia; Luppi, Mario; Torelli, Giuseppe
abstract
No abstract available
2003
- Post-transplant Kaposi sarcoma originates from the seeding of donor-derived progenitors
[Articolo su rivista]
Barozzi, Patrizia; Luppi, Mario; F., Facchetti; C., Mecucci; Alu, M.; R., Sarid; V., Rasini; L., Ravazzini; E., Rossi; S., Festa; B., Crescenzi; Dg, Wolf; Tf, Schulz; Torelli, Giuseppe
abstract
Kaposi sarcoma (KS) is a vascular tumor that can develop in recipients of solid tissue transplants as a result of either primary infection or reactivation of a gammaherpesvirus, the KS-associated herpesvirus, also known as human herpesvirus-8 (HHV-8). We studied whether HHV-8 and the elusive KS progenitor cells could be transmitted from the donor through the grafts. We used a variety of molecular, cytogenetic, immunohistochemical and immunofluorescence methods to show that the HHV-8-infected neoplastic cells in post-transplant KS from five of eight renal transplant patients harbored either genetic or antigenic markers of their matched donors. These data suggest the use of donor-derived HHV-8-specific T cells for the control of post-transplant KS.
2003
- Skin cancers after organ transplantation
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Torelli, Giuseppe
abstract
No abstract available
2002
- HHV-8 infection in the transplantation setting: A concern only for solid organ transplant patients?
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Rasini, Valeria; Torelli, Giuseppe
abstract
Early epidemiologic studies have shown an increased risk of Kaposi sarcoma (KS) in recipients of solid organ transplants, while KS is exceptional in the setting of autologous and allogeneic bone marrow (BM) and peripheral blood stem cell (PBSC) transplant patients. The recent discovery of human herpesvirus 8 (HHV-8) as the necessary etiologic agent of KS has stimulated studies to assess whether KS is the result of HHV-8 transmission from the donor or of reactivation of a pre-existing HHV-8 infection in the recipient host. An association of HHV-8 infection with lymphoid neoplasias has also been observed, identifying this herpesvirus as a possible causal agent of some Epstein-Barr virus negative post-transplant lymphoproliferative diseases. The recent description of non-neoplastic complications associated with HHV-8 reactivation after autologous PBSC transplantation, has raised concerns about the spectrum of diseases potentially associated with this herpesvirus, even in the setting of BM and or PBSC transplantation. The issue of HHV-8 transmission with the grafts, the problems inherent with the diagnosis and monitoring of active viral infection, the need for prevention and treatment of the clinical consequences of HHV-8 primary infection and reactivation cannot be underestimated, at least in areas endemic for HHV-8 infection.
2002
- Severe pancytopenia and hemophagocytosis after HHV-8 primary infection in a renal transplant patient successfully treated with foscarnet
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Rasini, V; Riva, Giovanni; Re, A; Rossi, Giorgio; Setti, G; Sandrini, S; Facchetti, F; Torelli, Giuseppe
abstract
We report the occurrence of human herpesvirus (HHV)-8 primary infection in an adult male kidney recipient. Four months after transplantation, the patient developed visceral Kaposi sarcoma, and 1 month later he presented with progressive and severe peripheral cytopenia, in the presence of a normocellular or hypercellular bone marrow (BM) with hemophagocytosis. HHV-8 was the sole pathogen detected by polymerase chain reaction either in the serum or in the BM. HHV-8 latent nuclear antigen was detected in immature progenitor cells from the BM. Immunosuppressive therapy was reduced, and the patient was treated with foscarnet for 2 weeks, leading to a dramatic normalization of blood cell counts, concomitantly with the disappearance of HHV-8 viremia. At the end of antiviral therapy, the patient received chemotherapy, and Kaposi sarcoma regressed in 2 months. Severe peripheral cytopenia may be a post-transplant complication after HHV-8 infection, for which treatment with foscarnet seems appropriate.
2001
- Angiogenesis in multiple myeloma: Correlation between in vitro endothelial colony growth (CFU-En) and clinical biological features
[Articolo su rivista]
Dominici, Massimo; Luppi, Mario; F., Lanza; Campioni, D.; Barozzi, Patrizia; S., Pauli; R., Milani; F., Cavazzini; M., Punturieri; R., Trovato; G., Torelli; G., Castoldi
abstract
Mouse models and studies performed on fixed bone marrow (BM) specimens obtained from patients with multiple myeloma (MM) suggest that plasma cell growth is dependent on endothelial cell (EC) proliferation within the BM microenvironment. In order to assess whether EC overgrowth in MM reflects a spontaneous in vitro angiogenesis, BM mononucleated cells from 13 untreated (UT) MM, 20 treated (11 with melphalan and nine with DAV schedule) MM, eight patients with monoclonal gammopathy of uncertain significance (MGUS) and eight controls were seeded in an unselective medium to assess EC proliferation. Furthermore, the influence of IL6 on the EC growth was investigated. Endothelial colonies (CFU-En) appeared as small clusters, formed by at least 100 slightly elongated and sometimes bi-nucleated cells expressing factor VIII, CD31 and CD105 (endoglin). The CFU-En mean number/10(6) BM mononucleated cells in untreated MM samples (2.07 s.d. +/- 1.3) was significantly higher than in normal BM (0.28 +/- 0.48), while no difference was seen between normal BM and MGUS (0.28 +/- 0.54). Interestingly, the mean number of CFU-En in the DAV group (1.88 +/- 1.6) did not differ from the UT, while it was found to be lower in the melphalan group (0.31 +/- 0.63). The addition of anti-IL6 monoclonal antibody induced a reduction of both the plasma cells in the supernatant and the CFU-En number. This study describes a rapid and feasible assay providing support for the association between EC and plasma cells further suggesting that the in vitro angiogenesis process may parallel that observed in vivo.
2001
- Immunoglobulin gene mutations and frequent use of VH1-69 and VH4-34 segments in hepatitis C virus-positive and hepatitis C virus-negative nodal marginal zone B-cell lymphoma
[Articolo su rivista]
Marasca, Roberto; Vaccari, Paola; Luppi, Mario; Zucchini, Patrizia; Castelli, Ilaria; Barozzi, Patrizia; A., Cuoghi; Torelli, Giuseppe
abstract
Nodal marginal zone B-cell lymphoma (NMZL) is actually considered as a distinct entity that must be distinguished from extra-nodal and splenic marginal zone Lymphomas. To define the cell origin and the role of antigen stimulation we determined the nucleotide sequence of the tumor-related immunoglobulin heavy chain variable genes in 10 cases of NMZL. The results were also evaluated on the basis of the presence of chronic hepatitis C virus (HCV) infection. All 10 cases harbored VH somatic mutations with a sequence homology compared to the closest germline gene, ranging from 83.33 to 98.28%. Interestingly, different VH segments were preferentially used in HCV-positive and HCV-negative patients: three of five HCV-negative NMZLs used a VH4-34 segment joined with different D and JH segments whereas three of five HCV-positive NMZLs used a VH1-69 gene joined with a D3-22 and a JH4 segment, with very strong similarities in the CDR3s among the three different cases. These data indicate: 1) NMZL is derived from B cells that have experienced the germinal center reaction; 2) the preferential usage of a VH1-69 segment in the majority of the HCV-positive NMZL cases with similar CDR3s suggests the presence of a common antigen, probably a HCV antigen epitope, involved in the B-cell selection; and 3) the use of a VH4-34 segment suggests a role of yet unknown B-cell superantigen(s) in the selection of tumor B-cell precursors in HCV-negative NMZL.
2000
- Bone marrow failure associated with human herpesvirus 8 infection after transplantation
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Schulz, Tf; Setti, G; Staskus, K; Trovato, R; Narni, Franco; Donelli, A; Maiorana, Antonino; Marasca, Roberto; Sandrini, S; Torelli, G.
abstract
BACKGROUND: Human herpesvirus 8 (HHV-8) infection has been linked to the development of Kaposi's sarcoma and to rare lymphoproliferative disorders.METHODS: We used molecular methods, serologic methods, in situ hybridization, and immunohistochemical analyses to study HHV-8 infection in association with nonmalignant illnesses in three patients after transplantation.RESULTS: Primary HHV-8 infections developed in two patients four months after each received a kidney from the same HHV-8-seropositive cadaveric donor. Seroconversion and viremia occurred coincidentally with disseminated Kaposi's sarcoma in one patient and with an acute syndrome of fever, splenomegaly, cytopenia, and marrow failure with plasmacytosis in the other patient. HHV-8 latent nuclear antigen was present in immature progenitor cells from the aplastic marrow of the latter patient. Identification of the highly variable K1 gene sequence of the HHV-8 genome in both the donor's peripheral-blood cells and the recipients' serum confirmed that transmission had occurred. HHV-8 viremia also occurred after autologous peripheral-blood stem-cell transplantation in an HHV-8-seropositive patient with non-Hodgkin's lymphoma. Reactivation of the infection was associated with the development of fever and marrow aplasia with plasmacytosis; there was no evidence of other infections. HHV-8 transcripts and latent nuclear antigen were expressed in the aplastic marrow but not in two normal marrow samples obtained before transplantation.CONCLUSIONS: Primary HHV-8 infection and reactivation of infection may be associated with nonneoplastic complications in immunosuppressed patients.
2000
- Detection of antibodies to human herpesvirus 8 in Italian children: evidence for horizontal transmission
[Articolo su rivista]
D., Whitby; Luppi, Mario; C., Sabin; Barozzi, Patrizia; AR Di, Biase; Balli, Fiorella; F., Cucci; Ra, Weiss; C., Boshoff; Torelli, Giuseppe
abstract
Human herpesvirus 8 (HHV-8), also known as Kaposi´s sarcoma associated herpesvirus (KSHV), has been shown to be the causative agent for Kaposi´s sarcoma (KS) and to be more prevalent in populations or risk groups at increased risk for KS. HHV-8 infection is rare in children from the US and the UK, but has been reported in African children. In this study we examine HHV-8 infection in children from Italy, a country with an elevated prevalence of HHV-8 in adults and high socio-economic conditions.
2000
- Fatal herpesvirus-6 encephalitis in a recipient of a T-cell-depleted peripheral blood stem cell transplant from a 3-loci mismatched related donor
[Articolo su rivista]
Tiacci, E; Luppi, Mario; Barozzi, Patrizia; Gurdo, G; Tabilio, A; Ballanti, S; Torelli, Giuseppe; Aversa, F.
abstract
Human herpesvirus-6 (HHV-6), like all the other herpes viruses, remains latent in host cells after primary infection but can be reactivated In immuno-compromised patients causing fever, skin rash, bone marrow (BM) suppression, pneumonitis, Sinusitis and meningoencephalitis. We describe the case of a man with chronic myelogenous leukemia who developed encephalitis associated with acute graft-versus-host disease two months after a T-cell-depleted mismatched peripheral blood stem cell transplant. Magnetic resonance images of the brain revealed multiple bilateral foci of signal abnormality. HHV-6 was the only pathogen detected in cerebrospinal fluid by PCR. treatment with both ganciclovir and foscarnet was unsuccessful and the patient gradually deteriorated and died. Other cases of HHV-6 encephalitis after bone marrow transplantation are reviewed.
2000
- Late-appearing PML/RAR alpha fusion transcript with coincidental t(12;13)(p13.2;q14) in acute promyelocytic leukemia lacking the t(15;17) cytogenetic anomaly
[Articolo su rivista]
Temperani, Paola; Luppi, Mario; F., Giacobbi; V., Medici; M., Morselli; Barozzi, Patrizia; Marasca, Roberto; Torelli, Giuseppe; Emilia, Giovanni
abstract
The late appearance of a cytogenetic/molecular hallmark in human leukemias is a rare event. We report on a case of acute myeloid leukemia with morphology, immunophenotype and clinical features typical of promyelocytic subtype (APL), in which the specific PML/RAR alpha gene rearrangement was molecularly detected only at second relapse of disease, without cytogenetic evidence of the t(15;17). The emergence of the PML/RAR alpha gene may be therapy-related or may represent the exceptional result of a clonal evolution during progression of neoplasia. At second relapse, a novel cell clone bearing a t(12;13)(p13.2;q14) was also observed and a molecular deletion and rearrangement of a locus at 13q14, distinct from retinoblastoma (Rb1) locus, was found. In this unusual case, the PML/RAR alpha product seems to be not essential for the expression of the promyelocytic phenotype at diagnosis and, when detectable, it is not the sole genetic defect. (C) 2000 Elsevier Science Inc. All rights reserved.
2000
- Molecular evidence of organ-related transmission of Kaposi sarcoma-associated herpesvirus or human herpesvirus-8 in transplant patients
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; G., Santagostino; R., Trovato; Tf, Schulz; Marasca, Roberto; D., Bottalico; L., Bignardi; Torelli, Giuseppe
abstract
In transplant patients, Kaposi sarcoma (KS)-associated herpesvirus or human herpesvirus-8 (HHV-8) infection is associated with the development of KS, primary effusion lymphoma and Castleman disease. Whether HHV-8 is either reactivated in the recipient or transmitted by the donor has been investigated so far only by serologic studies. Thus, we addressed the issue of HHV-8 transmission in the transplantation setting by molecular methods. We exploited the high level variability of the orf-K1 gene and the polymorphism of the orf-73 gene of the HHV-8 genome to assess the genetic relatedness of the HHV-8 strains identified in the posttransplant KS lesions that developed, simultaneously, 20 months after transplantation, in 2 recipients of twin kidneys from the same cadaver donor. The 100% identity of nucleotide sequence of the most variable viral region and the presence of the same, single orf-73 type in both patients provides strong molecular evidence of organ-related transmission of HHV-8 in the setting of transplantation.
2000
- Nonmalignant disease associated with human herpesvirus 8 reactivation in patients who have undergone autologous peripheral blood stem cell transplantation
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Tf, Schulz; R., Trovato; A., Donelli; Narni, Franco; J., Sheldon; Marasca, Roberto; Torelli, Giuseppe
abstract
Fever, cutaneous rash, and hepatitis-for which an infectious cause was suspected-developed in an Italian patient with non-Hodgkin lymphoma after autologous peripheral blood stem cell (PBSC) transplantation. Polymerase chain reaction (PCR) with degenerate primers for the highly conserved DNA polymerase gene of herpesviruses detected herpesvirus sequences 100% identical to human herpesvirus-8 (HHV-8) in serial cell-free serum samples, collected immediately before or concomitant with the occurrence of clinical symptoms; no other common infections were documented. The presence of the HHV-8 genome (clade C) was confirmed by PCR with HHV-8-specific primers for orf 26 and orf-K1, HHV-8 viremia was undetectable either before transplantation or when the patient was clinically asymptomatic. Semiquantitative PCR analysis showed variations of the viral load correlating with the clinical status. Anti-HHV-8 antibodies were detected before and after transplantation by an immunofluorescence assay for lytic antigens. Active HHV-8 infection may be associated with nonmalignant illness after PBSC/bone marrow transplantation. (Blood. 2000;96:2355-2357)
2000
- PCR with degenerate primers for highly conserved DNA polymerase gene of the herpesvirus family shows neither human herpesvirus 8 nor a related variant in bone marrow stromal cells from multiple myeloma patinets.
[Articolo su rivista]
Dominici, Massimo; Luppi, Mario; D., Campioni; F., Lanza; Barozzi, Patrizia; R., Milani; S., Moretti; G., Nadali; R., Spanedda; R., Trovato; Torelli, Giuseppe; G., Castoldi
abstract
The possibility has been raised that either a human herpesvirus-8 (HHV-8) variant or a novel, unidentified, gamma-herpesvirus related to HHV-8 is frequently associated with multiple myeloma (MM), which could explain the lack of antibodies to HHV-8 antigens and the discordant results from polymerase chain reaction (PCR) studies of HHV-8-specific sequences in MM patients. Thus, we used a sensitive PCR assay with degenerate primers targeting the highly conserved DNA polymerase gene of the herpesvirus family to examine the long-term cultures of bone marrow stromal cells (BMSCs) from 19 MM, 3 monoclonal gammopathies of undetermined significance and 6 control patients. Both the culture supernatant and the adherent stromal layer were examined from the 2nd until the 8th week of culture to assess the immunophenotype of the various cell types harvested for the molecular analysis. BMSCs consisted of a mixed population of fibroblast, macrophage, dendritic and endothelial cells. An amplified product of the expected size was obtained only in 3 MM cases, both in the adherent and nonadherent fractions. Direct sequencing and alignment of the nucleotide and amino acid sequences showed that the DNA sequences were 100% identical to Epstein-Barr virus (EBV) DNA. The PCR positivity was due to the presence of EBV-infected lymphoblastoid cells with plasmacytoid features, expressing the EBV-encoded latent membrane protein-1 and detectable either in the stromal cells or in the culture supernatant. Our data do not support a causal role of either HHV-8 or a novel herpesviral variant related to HHV-8 in MM.
1999
- Cellular localization of human herpesvirus 8 in nonneoplastic lymphadenopathies and chronic interstitial pneumonitis by in situ polymerase chain reaction studies
[Articolo su rivista]
Trovato, R; Luppi, Mario; Barozzi, Patrizia; Da Prato, L; Maiorana, Antonino; Lico, S; Marasca, Roberto; Torricelli, Pietro; Torelli, Giuseppe; Ceccherini Nelli, L.
abstract
Objectives: To study the cellular localization of human herpesvirus 8 (HHV-8) in rare cases of HHV-8 infection from Italy that are associated neither with human immunodeficiency virus (HIV) infection nor Kaposi's sarcoma (KS). Methods: The presence and distribution of HHV-8-infected cells was investigated by direct in situ polymerase chain reaction (PCR) in the lymph node tissues from 2 patients with reactive lymphadenopathies with florid follicular hyperplasia and increased vascularity and in the lung tissue from 1 patient with chronic interstitial pneumonitis. Results: HHV-8 was localized in lymphoid and monocyte-macrophage cells scattered in the interfollicular regions of both lymph nodes but not in endothelial cells. In the lung tissue, HHV-8 was found in the inflammatory cells infiltrating the interalveolar interstitium, in endothelial cells of the pulmonary vasculature, and in rare pneumocytes. Conclusions: HHV-8 can infect nonneoplastic lymph nodes of immunocompetent subjects, and the distribution of infected cells outside of the germinal centers resembles that of Epstein-Barr virus (EBV)-infected cells in the lymph nodes in the course of infectious mononucleosis. Endothelial cells and pneumocytes may be a target of HHV-8 infection out of the KS setting, at least in the presence of a chronic inflammatory process.
1999
- Erratum: Cellular localization of human herpesvirus 8 in nonneoplastic lymphadenopathies and chronic interstitial pneumonitis by in situ polymerase chain reaction studies (Journal of Human Virology (January/February 1999) 2:1 (38-44))
[Articolo su rivista]
Trovato, R.; Luppi, M.; Barozzi, P.; Da Prato, L.; Maiorana, A.; Lico, S.; Marasca, R.; Torricelli, P.; Torelli, G.; Ceccherini-Nelli, L.
abstract
1999
- Expression of cell-homologous genes of human herpesvirus-8 in human immunodeficiency virus-negative lymphoproliferative diseases
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Maiorana, Antonino; R., Trovato; Marasca, Roberto; M., Morselli; K., Cagossi; Torelli, Giuseppe
abstract
Human herpesvirus-8 (HHV-8) genome encodes for genes homologous to human cellular genes such as interleukin-6 (IL-6), Cyclin-D, BCL-5, and IL-8 receptor (G-protein-coupled receptor [GCR]), We used reverse transcriptase-polymerase chain reaction to study the expression of these viral genes in lymphoproliferative disorders associated with HHV-8 infection, None of these genes was expressed in 1 case of benign, localized Castleman´s disease (CD), and only viral IL-6 and viral Cyclin-D were transcribed in 2 cases of benign lymphadenopathies with giant germinal center hyperplasia and increased vascularity. In contrast, all 4 genes were transcribed in 1 case of multicentric CD of plasma cell type with aggressive clinical course and in 1 primary effusion lymphoma cell line. Our study provides the evidence that various HHV-8 genes, homologous to cellular genes involved in control of proliferation and apoptosis, may be differently expressed in different lymphoid disorders in vivo.
1999
- Fine mapping of an apparently targeted latent human herpesvirus type 6 integration site in chromosome band 17p13.3
[Articolo su rivista]
Morris, C; Luppi, Mario; Mcdonald, M; Barozzi, Patrizia; Torelli, Giuseppe
abstract
An unusually high level of latent HHV-6 infection has been documented in the peripheral blood and/or bone marrow cells of a small group of patients with predominantly malignant lymphoid disorders, and in at least one healthy individual, We have shown previously in peripheral blood mononuclear cells (PBMCs) of three patients, two with a history of lymphoma and one with multiple sclerosis, a specific target site for latent integration of the full-length HHV-6 viral genome on the distal short arm of chromosome 17, in band p13.3. Fluorescence in situ hybridization (FISH) procedures were used to map more precisely the location of the viral integration site in one of those patients, relative to two known oncogenes mapped previously, namely CRK, and the more telomeric ABR oncogene. It is shown that the HHV-6 integration site is located at least 1,000 kb telomeric of ABR, and is very likely to map close to or within the telomeric sequences of 17p. This finding is significant given that human telomeric-like repeats flank the terminal ends of the HHV-6 genome. Cytogenetic studies showed evidence of karyotype instability in the peripheral blood cells infected latently. J. Med. Virol. 58:69-75, 1999.
1999
- Human herpesvirus 6 latently infects early bone marrow progenitors in vivo
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; C., Morris; Maiorana, Antonino; R., Garber; G., Bonacorsi; A., Donelli; Marasca, Roberto; A., Tabilio; Torelli, Giuseppe
abstract
We have studied the in vivo tropism of human herpesvirus 6 (HHV-6) for hemopoietic cells in patients with latent HHV-6 infection. Having used a variety of cell purification, molecular, cytogenetic, and immunocytochemical procedures, we report the first evidence that HHV-6 latently infects early bone marrow progenitor cells and that HHV-6 may be transmitted longitudinally to cells which differentiate along the committed pathways.
1999
- Human herpesvirus-8 infection in hemodialysis patients from northern Italy.
[Articolo su rivista]
Luppi, Mario; Vandelli, L.; Whitby, D.; Savazzi, A. M.; Barozzi, Patrizia; Medici, G.; Albertazzi, Alberto; Torelli, Giuseppe
abstract
No abstract available
1999
- Lack of confirmation of an association between HTLV-I infection and myelodysplastic syndrome
[Articolo su rivista]
M., Morselli; Luppi, Mario; Barozzi, Patrizia; Dominici, Massimo; Temperani, Paola; D., Campione; F., Lanza; R., Trovato; Marasca, Roberto; G., Longo; G., Emilia; Torelli, Giuseppe
abstract
No abstract available
1999
- Neither Human Herpesvirus 8 nor a related variant could be detected in bone marrow stromal cells from multiple myeloma patients.
[Abstract in Rivista]
Dominici, Massimo; Luppi, Mario; D., Campioni; F., Lanza; Barozzi, Patrizia; R., Milani; A., Latorraca; S., Moretti; Torelli, Giuseppe; G. L., Castoldi
abstract
The possibility has been raised that either a human herpesvirus-
8 (HHV-8) variant or a novel, unidentified, g-herpesvirus
related to HHV-8 is frequently associated with multiple
myeloma (MM), which could explain the lack of antibodies to
HHV-8 antigens and the discordant results from polymerase
chain reaction (PCR) studies of HHV-8-specific sequences in
MM patients. Thus, we used a sensitive PCR assay with degenerate
primers targeting the highly conserved DNA polymerase
gene of the herpesvirus family to examine the longterm
cultures of bone marrow stromal cells (BMSCs) from
19 MM, 3 monoclonal gammopathies of undetermined significance
and 6 control patients. Both the culture supernatant
and the adherent stromal layer were examined from the 2nd
until the 8th week of culture to assess the immunophenotype
of the various cell types harvested for the molecular analysis.
BMSCs consisted of a mixed population of fibroblast, macrophage,
dendritic and endothelial cells. An amplified product
of the expected size was obtained only in 3 MM cases, both in
the adherent and nonadherent fractions. Direct sequencing
and alignment of the nucleotide and amino acid sequences
showed that the DNA sequences were 100% identical to
Epstein-Barr virus (EBV) DNA. The PCR positivity was due
to the presence of EBV-infected lymphoblastoid cells with
plasmacytoid features, expressing the EBV-encoded latent
membrane protein-1 and detectable either in the stromal
cells or in the culture supernatant. Our data do not support
a causal role of either HHV-8 or a novel herpesviral variant
related to HHV-8 in MM.
1999
- PCR con primer degenerati non evidenzia sequenze virali di HHV-8 ne di una sua variante in cellule stromali midollari di pazienti affetti da gammopatie monoclonali
[Abstract in Rivista]
Dominici, Massimo; Luppi, Mario; D., Campioni; F., Lanza; Barozzi, Patrizia; R., Milani; S., Moretti; R., Spanedda; Torelli, Giuseppe; G. L., Castoldi
abstract
The association of human herpesvirus-8 (HHV-8) infection with multiple myeloma (MM) is still controversial. The possibility has been raised that either a HHV-8 variant or a novel, as yet unidentified, γ-herpesvirus related to HHV-8, are frequently associated with MM, which could explain the lack of antibodies to HHV-8 antigens and the discordant results from polymerase chain reaction studies (PCR) of HHV-8 specific genomic sequences in MM patients. Thus, we used a sensitive PCR assay with degenerate primers targeting the highly conserved DNA polymerase gene of the herpesvirus family to examine the long term-cultures (LTC) of bone marrow stromal cells (BMSCs) from 21 MM, 3 monoclonal gammopathy of undetermined significance (MGUS) and 6 control patients. Both the culture supernatant and the adherent stromal layer were examined from the 2nd until the 8th week of culture to assess the immunophenotype of the various cell types harvested for the molecular analysis. BMSCs consisted of a mixed population of fibroblast, macrophage, dendritic and endothelial cells. An amplified product of the expected size was obtained only in 3 MM cases, both in the adherent and non adherent fractions. Direct sequencing and alignment of the nucleotide and amino acidic sequences showed that the DNA sequences were 100% identical to Epstein-Barr virus DNA. PCR with specific primers targeting the latent membrane protein-1 (LMP-1) gene confirmed the presence of EBV. The PCR positivity was due to the presence of EBV infected lymphoblastoid cells (LCLs) with plasmacytoid features, expressing the LMP-1 protein and detectable either in the stromal cells or in the culture supernatant. Our data do not support a causal role of either HHV-8 or a novel herpesviral variant related to HHV-8 in MM, not even in the presence of EBV co-infection.
1999
- The oncogenic 30 and 69 bp deletion variants of the EBV LMP-1 gene are common in HIV-negative lymphoproliferations, both malignant and benign
[Articolo su rivista]
Barozzi, Patrizia; Luppi, Mario; K., Cagossi; Maiorana, Antonino; Marasca, Roberto; T., Artusi; S., Poggi; S. A., Pileri; Torelli, Giuseppe
abstract
BACKGROUND:
In vitro studies have shown that the 30 and 69 base pair (bp) deletion variants of the latent membrane protein (LMP)-1 gene of the Epstein-Barr virus (EBV) have a higher transforming capacity than the wild-type variant. In recent years these studies have triggered an in vivo search for such potentially oncogenic variants in lymphoid tissues.
PATIENTS AND METHODS:
We used polymerase chain reaction (PCR) to investigate the prevalence of LMP-1 gene variants in EBV-positive lymph nodes from 60 HIV-negative Italian patients with benign and malignant lymphoid disorders.
RESULTS:
The 30 bp variant was detected in 10 of 39 (25.6%) malignant lymphomas but also in 4 of 13 (30%) reactive lymphadenitis with follicular hyperplasia. Of note is the fact that the 69 bp variant was detected in three cases of malignant lymphoproliferation but also in two cases of localized Castleman's disease of hyalin vascular type.
CONCLUSIONS:
The molecular detection of the oncogenic variants of the LMP-1 gene in a lymph node biopsy as an indicator of the aggressiveness of the EBV-associated lymphoproliferative disease must be considered with caution. The relatively high frequency of the 69 bp variant in our series compared with that reported in the literature probably reflects a different incidence of LMP-1 variants in healthy populations from different geographical areas.
1998
- Age- and sex-specific seroprevalence of human herpesvirus 8 in Jamaica - Response
[Articolo su rivista]
Luppi, Mario; Whitby, D; Barozzi, Patrizia; Boshoff, C; Weiss, R; Cucci, F; Torelli, Giuseppe
abstract
No abstract available
1998
- Age- and sex-specific seroprevalence of human herpesvirus 8 in Jamaica [3] (multiple letters)
[Articolo su rivista]
Manns, A.; Strickler, H. D.; Hanchard, B.; Manassaram, D. M.; Waters, D.; Ablashi, D. V.; Luppi, M.; Whitby, D.; Barozzi, P.; Boshoff, C.; Weiss, R.; Cucci, F.; Torelli, G.
abstract
1998
- Clinico-pathological characterization of hepatitis C virus-related B-cell non-Hodgkin's lymphomas without symptomatic cryoglobulinemia
[Articolo su rivista]
Luppi, Mario; G., Longo; Mg, Ferrari; Barozzi, Patrizia; Marasca, Roberto; M., Morselli; C., Valenti; Mascia, Maria Teresa; L., Vandelli; D., Vallisa; L., Cavanna; Torelli, Giuseppe
abstract
Background. Epidemiological evidence has suggested an association between hepatitis C virus (HCV) infection and B-cell lymphoproliferation. We studied the prevalence of HCV infection in a series of de novo B-cell non-Hodgkin's lymphoma (B-NHL) cases and correlated virological findings with clinico-histological features. Patients and methods. One hundred fifty-seven patients with de novo B-NHL were included in the study. Their serum was examined by ELISA and RIBA for the presence of anti-HCV antibodies, and either the peripheral blood mononuclear cells or the pathology tissues of all of the patients were examined by reverse transcriptase polymerase chain reaction for the presence of HCV RNA sequences, Results: HCV infection occurred in 22.3% of B-NHL patients and was documented before the diagnosis in about halfof the positive cases. Of interest. HCV infection was more frequently found in follicular center; marginal zone and diffuse large-cell lymphoma types, but was not associated with symptomatic cryoglobulinemia. The median survival time was 48 months in HCV-positive and 52 months in HCV-negative B-NHL patients. Conclusions. Our findings strengthen the pathogenetic link between HCV and B-NHL and show that HCV infection may be associated with the malignant proliferation of defined B-cell subsets other than the immunoglobulin Mk B-cell subset involved in the pathogenesis of mixed cryoglobulinemia type II and associated lymphoplasmacytoid lymphoma type. HCV-related liver disease did not affect the survival of our B-NHL patients.
1998
- Expression of human herpesvirus-6 antigens in benign and malignant lymphoproliferative diseases
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; R., Garber; Maiorana, Antonino; G., Bonacorsi; T., Artusi; R., Trovato; Marasca, Roberto; Torelli, Giuseppe
abstract
Immunohistochemistry was used to look for the expression of human herpesvirus-6 (HHV-6) antigens in a well characterized series of benign, atypical, and malignant lymphoid lesions, which tested positive for the presence of HHV-6 DNA, A panel of specific antibodies against HHV-6 antigens, characteristic either of the early (p41) or late (p101K, gp106, and gp116) phases of the viral cycle, was applied to the lymphoid tissues from 15 non-Hodgkin's lymphomas, 14 Hodgkin's disease cases, 5 angioimmunoblastic lymphadenopathies with dysproteinemia, 14 Reactive lymphadenopathies, and 2 cases of sinus histiocytosis with massive lymphadenopathy (Rosai-Dorfman disease). In lymphomatous tissues, the expression of late antigens was documented only in reactive cells, and mainly in plasma cells. Of interest, the expression of the early p41 antigen was detected in the so-called mummified Reed-Sternberg cells, in two Hodgkin's disease cases. In reactive lymphadenopathies, the HHV-6 late antigen-expressing cells were plasma cells, histiocytes, and rare granulocytes distributed in interfollicular areas. In both cases of Rosai-Dorfman disease, the p101K showed an intense staining in follicular dendritic cells of germinal centers, whereas the gp106 exhibited an intense cytoplasmic reaction in the abnormal histiocytes, which represent the histological hallmark of the disease. The expression of HHV-6 antigens is tightly controlled in lymphoid tissues. The lack of HHV-6 antigen expression in neoplastic cells and the limited expression in degenerating Reed-Sternberg cells argue against a major pathogenetic role of the vines in human lymphomagenesis, The detection of a rather unique pattern of viral late antigen expression in Rosai-Dorfman disease suggests a possible pathogenetic involvement of HHV-6 in some cases of this rare lymphoproliferative disorder.
1998
- Human herpesvirus 8 seroprevalence in blood donors and lymphoma patients from different regions of Italy
[Articolo su rivista]
D., Whitby; Luppi, Mario; Barozzi, Patrizia; C., Boshoff; Ra, Weiss; Torelli, Giuseppe
abstract
No abstract available
1998
- Integration of human herpesvirus 6 genome in human chromosomes
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Morris, Cm; Merelli, E; Torelli, Giuseppe
abstract
No abstract available
1998
- Polymerase chain reaction detection of human herpesvirus 8 sequences in primary central nervous system lymphomas
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Marasca, Roberto; Savarino, M; Torelli, Giuseppe
abstract
No abstract available
1997
- Hepatitis C virus-induced leuko-thrombocytopenia and haemolysis
[Articolo su rivista]
Emilia, G; Luppi, Mario; Ferrari, Mg; Barozzi, Patrizia; Marasca, Roberto; Torelli, Giuseppe
abstract
Hepatitis C virus (HCV) has been recognized as the cause of thrombocytopenia occurring in patients with chronic hepatitis C, possibly through autoimmune mechanisms. A patient is described with B cell chronic lymphocytic leukaemia, presenting with a marked leuko-thrombocytopenia and an associated mild haemolysis secondary to HCV infection, in the absence of clinical and biochemical signs of hepatitis. Anti-HCV antibodies were detected in the serum both by ELISA and RIBA but not 2 months before the onset of cytopenia. The presence of HCV RNA was documented both in the peripheral blood mononuclear cells and in the bone marrow by reverse transcriptase polymerase chain reaction of the 5´ untranslated region of the viral genome. Of interest, HCV RNA was also found in the serum, showing that viraemia was associated with the presence of circulating anti-HCV antibodies. HCV genotyping, performed by PCR amplification of the core region, revealed the presence of an unclassifiable genotype. The hypothetical mechanisms leading to HCV-induced cytopenia in this patient are briefly discussed. Treatment with corticosteroids was effective in controlling blood cell counts, without increasing viraemia and deterioration of liver disease. HCV infection should be considered in the differential diagnosis of possible causes of cytopenia, mainly in immunosuppressed patients, even in absence of clinical and biochemical signs of hepatitis.
1997
- Human Herpesvirus 8 strain variability in clinical conditions other than Kaposi’s sarcoma
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Marasca, Roberto; Ferrari, Mg; Torelli, Giuseppe
abstract
No abstract available
1997
- Human herpesvirus-6 reactivation in a longitudinal study of two HIV-1 infected patients
[Articolo su rivista]
Iuliano, R; Trovato, R; Lico, S; Luppi, Mario; Forastieri, G; Barsanti, La; Pizzigallo, Am; Mecocci, L; Barozzi, Patrizia; Torelli, Giuseppe; Mazzotta, F; Ceccherininelli, L.
abstract
After primary infection, human herpesvirus-6 (HHV-6) persists in latent form and can be reactivated in immunocompromised subjects. A longitudinal study of HHV-6 infection was carried out in two HIV-1 seropositive patients to provide in vivo evidence of HHV-6 reactivation. Concomitant with a significant rise of anti-HHV-6 IgG detected by IFA, a transient increase of HHV-6 viral load was shown in PBLs by PCR. During HHV-6 reactivation it was also identified either cell-free HHV-6 by PCR in plasma or IgM antibody titers. HHV-6 reactivation was followed by a temporary decrease in CD4+ count and by a progressive dramatic loss of CD4+ during the following 18 months. HHV-6 strain characterization by PCR demonstrated that first patient (MM) initially showed the B variant, followed by reactivation and persistence of the A variant, while in the second (SG) only the A variant was detected. The evidence of HHV-6 reactivation suggests its involvement in immunologic damage underlying the disease.
1997
- Prevalence of HCV infection and second neoplasms in marginal zone lymphomas
[Articolo su rivista]
Luppi, Mario; Longo, G; Ferrari, Mg; Ferrara, L; Marasca, Roberto; Barozzi, Patrizia; Morselli, M; Emilia, G; Torelli, Giuseppe
abstract
No abstract available
1996
- Additional neoplasms and HCV infection in low-grade lymphoma of MALT type
[Articolo su rivista]
Luppi, Mario; Longo, G; Ferrari, Mg; Ferrara, L; Marasca, Roberto; Barozzi, Patrizia; Morselli, M; Emilia, G; Torelli, Giuseppe
abstract
Several chronic inflammatory conditions and genetic alterations are likely to be involved in the pathogenesis of low-grade lymphoma of MALT type. In a well-characterized series of 27 patients with low-grade lymphoma of MALT type, we studied: (1) the incidence of other neoplasms, which might be indicative of genetic instability, apparently a characteristic of this disease; (2) the prevalence of serologic and molecular markers of HCV infection, which has been found in association with other lymphoproliferative disorders. Three patients had one or more additional cancers; a total of eight tumours, five of which occurred in the same patient, suggests the presence of some genetic instability in at least some cases of the disease. Rather unexpectedly, anti-HCV antibodies and HCV RNA sequences were documented in 50% of the patients examined, without elevation of serum transaminases. Of interest, the two patients with parotid and conjunctival MALT lymphomas, respectively, with a previous history of Sjogren's syndrome, were HCV positive. We suggest, for the first time, that HCV may be considered, in addition to Helicobacter pylori, as another potential infectious co-factor in the multistep pathogenesis of low-grade lymphomas of MALT type.
1996
- Frequency and distribution of herpesvirus-like DNA sequences (KSHV) in different stages of classic Kaposi's sarcoma and in normal tissues from Italian population.
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Maiorana, Antonino; G., Collina; M. G., Ferrari; Marasca, Roberto; M., Morselli; E., Rossi; L., Ceccherini Nelli; Torelli, Giuseppe
abstract
The frequency and distribution of herpesvirus-like DNA sequences (KSHV) were investigated by PCR in the pathologic skin lesions of a series of 22 HIV-negative elderly patients with classic Kaposi´s sarcoma (KS) from Italy, one of the few regions of the world where classic KS is prevalent. Viral sequences were clearly identifiable in 15 cases, in particular in 2 of 5 patch, in 3 of 6 plaque and in 10 of 11 nodular lesions. Our findings confirm the association of these herpesvirus-like DNA sequences with KS in unrelated populations, providing evidence of the putative KS-associated agent in all different histologic lesions of the disease, mainly in the nodular stage. The search for other herpesviruses by PCR showed that Epstein-Barr virus (EBV) sequences were present in 7 of 22 pathologic skin lesions. In 4 cases, both EBV and KSHV were present. On the contrary, all 22 classic KS specimens were negative for human herpesvirus-6 sequences. Two of 3 patch and the 1 nodular lesions from AIDS-related KS patients examined were positive for KSHV but negative for both EBV and HHV-6 sequences. Furthermore, we evaluated the prevalence of KSHV sequences in the normal population of the same geographical area. Thirteen peripheral blood mononuclear cell samples, 9 salivary gland tissues and 6 saliva samples from healthy subjects were invariably found negative for KSHV, using the same PCR technique. Of interest, 2 of 11 hyperplastic tonsils harboured these herpesvirus-like sequences, suggesting that, like other herpesviruses, the KS-associated agent may be harboured in a proportion of normal individuals and tonsils may represent at least one of the possible reservoirs of this putative lymphotropic gamma-herpesvirus in vivo.
1996
- HHV-8-associated primary cerebral B-cell lymphoma in HIV-negative patient after long-term steroids
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Marasca, Roberto; Savarino, M; Torelli, Giuseppe
abstract
No abstract available
1996
- Hepatitis C virus infection in subsets of neoplastic lymphoproliferations not associated with cryoglobulinemia
[Articolo su rivista]
Luppi, Mario; Ferrari, Mg; Bonaccorsi, G; Longo, G; Narni, Franco; Barozzi, Patrizia; Marasca, Roberto; Mussini, C; Torelli, Giuseppe
abstract
Hepatitis C virus (HCV) is both hepatotropic and lymphotropic and a dear-cut association has been proposed between HCV infection and mixed cryoglobulinemia (MC), a benign lymphoproliferative disorder, which sometimes evolves into a frank malignant B cell non-Hodgkin's lymphoma (B-NHL). Moreover, the presence of antibodies to HCV, as well as of HCV-specific genomes has been reported in the sera of over 37% patients with B-NHL, not associated with MC. Thus, we decided to perform both a serologic and a molecular study to give insights into a possible relationship between HCV infection and neoplastic lymphoproliferations. We used ELISA and RIBA tests to show that anti-HCV antibodies were present in the serum of 29 out of 69 unselected B-NHL patients (42%), while seropositivity in a healthy population was about 1%. The prevalence of anti-HCV antibodies was low in definite subsets of B lymphoid disorders, including multiple myeloma, Waldenstrom's macroglobulinemia and monoclonal gammopathies of undetermined significance. Then, using reverse transcriptase polymerase chain reaction, we detected HCV sequences directly in the pathologic lymph node biopsies in 13 out of 34 B-NHL cases, and in particular in six out of eight low-grade lymphomas of MALT type and in five out of eight centroblastic-centrocytic follicular lymphomas. In contrast, the peripheral blood samples from 10 B cell chronic lymphocytic leukemia patients resulted negative for the presence of HCV genomes. Similarly, viral sequences were absent in 10 T cell NHL, while only one out of the 14 Hodgkin's disease cases tested resulted positive. Finally, we used a PCR-based assay to characterize the genotypes (I-IV) present in the positive lymphomatous tissues. The presence of both serologic and molecular markers of HCV infection in a high percentage of certain types of B-NHL, not associated with cryoglobulinemia, and its absence from other lymphoproliferative diseases extends the spectrum of HCV-associated lymphoproliferations arguing in favor of some role of this viral infection in the pathogenesis of the malignant proliferation of definite B lymphoid populations.
1996
- Human herpesvirus-8 DNA sequences in Human Immunodeficiency Virus-negative angioimmunoblastic lymphadenopathy and benign lymphadenopathy with giant germinal center hyperplasia and increased vascularity
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Maiorana, Antonino; T., Artusi; R., Trovato; Marasca, Roberto; M., Savarino; L., Ceccherini Nelli; Torelli, Giuseppe
abstract
Human herpesvirus-8 (HHV-8) DNA sequences have been reported to be strictly associated not only with various forms of Kaposi´s sarcoma but also with an unusual subgroup of acquired immunodeficiency syndrome (AIDS)-related B-cell lymphomas. A possible relation of this putative virus also with multicentric Castleman´s disease (MCD) has been recently suggested. We used polymerase chain reaction to look for the presence of HHV-8 sequences in a well characterized series of benign, atypical, and malignant lymphoid tissues from 45 Hodgkin´s disease and 43 non-Hodgkin´s lymphoma (NHL) cases, as well as from 5 MCD, 15 angioimmunoblastic lymphadenopathy (AILD), and 23 benign lymphadenopathy cases. Among the 38 AIDS-related lymphoid lesions, only 1 NHL and 1 persistent generalized lymphadenopathy (PGL) case were positive. Furthermore, among the 92 non-AIDS-related lymphoproliferative disorders, HHV-8 sequences were detected in 3 classic AILD cases and in 4 reactive lymphadenopathies. Six of 9 HHV-8 positive lymphoid lesions (1 NHL, 1 PGL, 1 AILD, and 3 reactive lymphadenopathy cases) were also positive for Epstein-Barr viral sequences. The four human immunodeficiency virus (HIV) negative lymphadenopathies positive for HHV-8 sequences showed an almost identical histology, characterized by a predominantly follicular lesion, with giant germinal center hyperplasia, and increased vascularity, resembling HIV-related lymphadenopathy and MCD. Our results, while providing the first evidence of the presence of HHV-8 sequences in AILD cases, suggest a possible association of these herpesviral sequences with a distinct hystologic type of non-neoplastic lymphadenopathy, not associated with other common herpes infections.
1996
- Lymphotropic herpes virus (EBV, HHV-6, HHV-8) DNA sequences in HIV negative Castleman's disease
[Articolo su rivista]
Barozzi, Patrizia; Luppi, Mario; Masini, L; Marasca, Roberto; Savarino, M; Morselli, M; Ferrari, Mg; Bevini, M; Bonacorsi, G; Torelli, Giuseppe
abstract
Aim-To evaluate the possible involvement of lymphotropic herpes viruses in Castleman's disease. Methods-Archival formalin fixed, paraffin wax embedded biopsy specimens from 16 HIV negative patients (11 with localised and five of multicentric disease) were studied. Epstein-Barr virus (EBV), human herpes virus-6 (HHV-6) and human herpes virus-8 (HHV-8) DNA was detected using PCR. PCR was also used to characterise the EBV genomes and the clonal status of the lesions. Results-EBV sequences were identified in nine (56%) cases. The main EBV genotype detected was type 1. Two (12%) cases were positive for both HHV-6 and EBV sequences. HHV-8 sequences were detected in one case of localised Castleman's disease, the sequence of which differed from that of the HHV-8 prototype. No clonal immunoglobulin gene rearrangements were found. Conclusions-EBV DNA was detected in a substantial proportion of cases, suggesting that it may have a role in the pathogenesis of Castleman's disease, unlike HHV-6 which was detected rarely. This is the first report of HHV-8 specific sequences in the localised from of the disease.
1996
- P53 gene mutations in chronic myelogenous leukemia medullary and extramedullary blast crisis
[Articolo su rivista]
Marasca, Roberto; Luppi, Mario; Barozzi, Patrizia; Ferrari, Mg; Morselli, M; Torelli, Giuseppe
abstract
Molecular alterations of the P53 gene were investigated in 27 unselected patients with chronic myelogenous leukemia (CML) blast crisis. A rearrangement of the P53 gene was evident by Southern blotting in 3 cases, one of which also showed the same alteration in the chronic phase. Single strand conformation polymorphism and sequencing analysis showed point mutations in 4 blast crisis cases. Of interest, P53 point mutations were evident in all the 3 cases of extramedullary blast crisis examined and the same point mutation was found in the myeloblastoma tissues and in the subsequent peripheral blast cells. These data indicate that: a) P53 gene mutations occur in a significant but not a large number of CML acute phase cases; b) P53 gene point mutations seem to correlate strongly with the infrequent extramedullary presentation of the blast crisis; c) the presence of the same P53 gene point mutation in extramedullary and bone marrow blast cells confirms the common clonal origin of the two cellular populations.
1995
- HUMAN HERPESVIRUS-6 - A SURVEY OF PRESENCE AND DISTRIBUTION OF GENOMIC SEQUENCES IN NORMAL BRAIN AND NEUROGLIAL TUMORS
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Maiorana, Antonino; Marasca, Roberto; R., Trovato; Fano, Rita Adriana; L., CECCHERINI NELLI; Torelli, Giuseppe
abstract
In an attempt to study the frequency and distribution of human herpesvirus-6 (HHV-6) infection both in normal and neoplastic brain tissues in vivo, polymerase chain reaction was used to look for HHV-6 genomes: 1) in samples, obtained at necropsy, from different regions of the brain of immunocompetent adult subjects and of patients who died of AIDS; 2) in the surgical biopsies of a well-characterized series of primary brain tumors of neuroglial origin. HHV-6-specific sequences were identified in six of nine brain samples from immunocompetent subjects, and in four of seven brain samples from AIDS patients. Viral sequences were identified in the specimens derived either from the grey (frontal cortex and basal ganglia) or from the periventricular white matter. HHV-6 DNA was found only in 6 of the 37 primary brain tumor biopsies examined. Th is study provides for the first time molecular evidence of a wide distribution of HHV-6 infection in the brain tissues of a high proportion of subjects, both in normal and in impaired immunity. in this large series of tumor biopsies the presence of HHV-6 genomic sequences is a rare phenomenon, arguing against a major role of this herpesvirus in the pathogenesis of primary brain tumors of neuroglial origin in immuno-competent subjects.
1995
- Human herpesvirus-6 (HHV-6) in blood donors
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Marasca, Roberto; Ceccherini Nelli, L; Ceccherelli, G; Torelli, G.
abstract
No abstract available
1995
- Human herpesvirus-6 genome in acute lymphoblastic leukemia: Evidence against an etiologic relationship
[Articolo su rivista]
Barozzi, Patrizia; Luppi, Mario; Marasca, Roberto; Trovato, R; Ceccherininelli, L; Torelli, Giuseppe
abstract
No abstract available
1995
- Spontaneous loss of Ph chromosome with maintenance of clonal hemopoiesis in an untreated patient with myeloproliferative disease and a long survival.
[Articolo su rivista]
Luppi, Mario; Morselli, M; Emilia, G; Temperani, P; Marasca, Roberto; Barozzi, Patrizia; Selleri, L; Torelli, G.
abstract
The unusual case of myeloproliferative disease described here is characterized by the following features: (1) a clinically completely silent course for 11 years without splenomegaly, marrow fibrosis, or cellular morphologic alterations; (2) the presence, at the onset, of a Philadelphia (Ph) chromosome without DNA breakpoints in the M-bcr region; (3) the spontaneous loss of detectable Ph-positive cells, 5 years after the first finding of leukocytosis, in the absence of any therapy; (4) the maintenance of the clonal nature of hematopoiesis, as revealed by the PGK X-linked inactivation pattern, in the absence of the Ph chromosome; and (5) a biphasic trend in the levels of leukocytes, red cells, and platelets during the years of observation.
1995
- Targeted integration of human herpesvirus 6 in the p arm of chromosome 17 of human peripheral blood mononuclear cells in vivo
[Articolo su rivista]
Torelli, Giuseppe; Marasca, Roberto; Paola, Cocconcelli; Elisa, Merelli; Luca Ceccherini, Nelli; Ferrari, Sergio; Mario, Luppi; Barozzi, Patrizia
abstract
Out of 64 cases of non-Hodgkin's lymphomas (NHL), 55 cases of Hodgkin's disease (HD) and 31 cases of multiple sclerosis (MS), 2 NHL, 7 HD and 1 MS cases were found positive by polymerase chain reaction (PCR) for the presence of HHV-6 sequences in pathologic lymph nodes of the lymphomas and in peripheral blood mononuclear cells (PBMCs) of MS. A further analysis of the PBMCs of the PCR positive cases by standard Southern blot technique revealed only 2 NHL, 3 HD and 1 MS cases as positive, indicating that these six patients have an unusually high viral copy number in the PBMCs. Restriction analysis, carried out using probes representative of different regions of the virus, showed that three cases retain only a deleted portion of the viral genome. In the remaining three cases a complete viral genome was present, containing the right end sequences in which the rep-like gene, possibly crucial to the viral and cellular life cycle, is located. The analysis by pulsed field gel electrophoresis (PFGE) of the total DNA of the PBMCs obtained directly, without culture from PBMCs of these last three cases (1 NHL, 1 HD, and 1 MS), using the same probes, showed the absence of free viral molecules and the association of viral sequences with high molecular weight DNA. These results are consistent with in vivo integration of the entire virus in the cellular genome. A further study of the same patients with chromosome fluorescence in situ hybridization (FISH) showed in all the three cases the presence of a specific hybridization site, located at the telomeric extremity of the short arm of chromosome 17 (17p13), suggesting that this location is at least a preferred site of an infrequent, but possibly biologically important, integration phenomenon.
1994
- Clonal nature of hypereosinophilic syndrome
[Articolo su rivista]
Luppi, Mario; Marasca, Roberto; Morselli, M; Barozzi, Patrizia; Torelli, Giuseppe
abstract
No abstract available
1994
- DOUBLE P53 POINT MUTATION IN EXTRAMEDULLARY BLAST CRISIS OF CHRONIC MYELOGENOUS LEUKEMIA
[Articolo su rivista]
Marasca, Roberto; Longo, G; Luppi, Mario; Barozzi, Patrizia; Torelli, Giuseppe
abstract
A patient with Philadelphia-positive chronic myelogenous leukemia (CML) evolved in extra-medullary blast crisis, was studied for the presence of alterations of the P53 tumor suppressor gene in the different stages of disease progression. No P53 gene aberrations were detected during the chronic and accelerated phases. Two identical missense point mutations, involving codons 249 and 281 and leading to the amino acid substitutions Arg-Ser and Thy-Asp, were identified in cells of an extramedullary mass and then in peripheral blood blast crisis cells. The data indicate that the medullary and extramedullary blast cells belong to the same cellular clone. They also strongly suggest that in this case, the alteration of P53 gene is strictly related to the progression of the disease, although this mechanism is certainly neither the only nor the most frequent molecular event leading to the acute phase.
1994
- Human herpesvirus 6 infection in normal human brain tissue
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Maiorana, Antonino; Marasca, Roberto; Torelli, Giuseppe
abstract
No abstract available
1994
- INTEGRATION OF HUMAN HERPESVIRUS-6 (HHV-6) GENOME IN CHROMOSOME-17 IN 2 LYMPHOMA PATIENTS
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Marasca, Roberto; Torelli, Giuseppe
abstract
No abstract available
1993
- 3 CASES OF HUMAN HERPESVIRUS-6 LATENT INFECTION - INTEGRATION OF VIRAL GENOME IN PERIPHERAL-BLOOD MONONUCLEAR CELL-DNA
[Articolo su rivista]
Luppi, Mario; Marasca, Roberto; Barozzi, Patrizia; Ferrari, Sergio; Ceccherininelli, L; Batoni, G; Merelli, E; Torelli, Giuseppe
abstract
Saliva and peripheral blood mononuclear cells from three patients, two with lymphoproliferative disorders and one suffering from multiple sclerosis, were examined for the presence of human herpesvirus-6 (HHV-6) genome by using the polymerase chain reaction and Southern blot analysis. The search for anti-HHV-6 antibodies, carried out in the sera of the same cases by an immunofluorescence assay, was negative in two cases at the lowest dilution used (1:40). These three patients had a high number of HHV-6 specific sequences in uncultured peripheral blood mononuclear cells, which are thought to be a normal site of viral latency although, in healthy individuals, the infected cells are extremely rare. In order to gain some insight into the state of the viral genome in this latent HHV-6 infection, we used pulsed field gel electrophoresis to separate HHV-6 DNA directly from HHV-6 (strain GS) infected HSB-2 cells and from the peripheral blood mononuclear cells of these three patients. Our study showed the presence of intact viral genome, of the expected length of 170 kb, persisting as free extrachromosomal element in the HSB-2 cells but not in patients' peripheral blood mononuclear cells. On the other hand, in strong contrast with the results obtained in infected HSB-2 DNA, the restriction analysis of the three patients' peripheral blood mononuclear cell DNA showed fragments of molecular weight constantly higher than the 170 kb segment, indicating that the viral sequences are linked to high molecular weight cellular DNA. Our findings are consistent only with a latent infection in which HHV-6 is integrated in vivo and suggest that pulsed field gel electrophoresis analysis is well worth using to evaluate the presence of integrated, intact, or fragmented viral genomes in HHV-6 associated lymphoproliferative diseases and immune disorders.
1993
- CHARACTERIZATION OF HUMAN HERPESVIRUS-6 GENOMES FROM CASES OF LATENT INFECTION IN HUMAN LYMPHOMAS AND IMMUNE DISORDERS
[Articolo su rivista]
Luppi, Mario; Barozzi, Patrizia; Marasca, Roberto; Ceccherininelli, L; Torelli, Giuseppe
abstract
No abstract available
1993
- FREQUENT DETECTION OF HUMAN HERPESVIRUS-6 SEQUENCES BY POLYMERASE CHAIN-REACTION IN PARAFFIN-EMBEDDED LYMPH-NODES FROM PATIENTS WITH ANGIOIMMUNOBLASTIC LYMPHADENOPATHY AND ANGIOIMMUNOBLASTIC LYMPHADENOPATHY-LIKE LYMPHOMA
[Articolo su rivista]
Luppi, Mario; Marasca, Roberto; Barozzi, Patrizia; Artusi, T; Torelli, Giuseppe
abstract
In search of a possible involvement of viral agents in angioimmunoblastic lymphadenopathy with dysproteinaemia (AILD) and AILD-like lymphoma (AILD-L), we studied the presence of human herpesvirus-6 (HHV-6) in the lymph node biopsies of 12 cases by polymerase chain reaction (PCR). Given the rarity of this lymphoproliferative disorder, we investigated archival specimens, consisting of formalin-fixed and paraffin-embedded tissues, obtained from patients with a clinical and histologic diagnosis of AILD and AILD-L. HHV-6 sequences were detected in the lymph node biopsies of 7 out of the 12 AILD and AILD-L cases examined. HHV-6 sequences were identified also in the involved liver and spleen tissues of one patient and in the PBMCs of two patients, all carrying viral sequences in the affected lymph nodes. We also used PCR to characterize the HHV-6 genomes, showing that two different viral strains are represented in the pathologic tissues. This study provides evidence of the presence of HHV-6 specific sequences in an unexpectedly high proportion of our series of AILD and AILD-L cases, suggesting a possible involvement of this lymphotropic virus in the pathogenesis of at least some cases of the disease.
1992
- HUMAN HERPESVIRUS-6 IN NON-AIDS RELATED HODGKINS AND NON-HODGKINS-LYMPHOMAS
[Articolo su rivista]
Torelli, Giuseppe; Marasca, Roberto; Montorsi, M; Luppi, Mario; Barozzi, Patrizia; Ceccherini, L; Batoni, G; Bendinelli, M; Muyombano, A.
abstract
Human herpesvirus 6 in non-AIDS related Hodgkin's and non-Hodgkin's lymphomas.