[Show abstract][Hide abstract] ABSTRACT: According to the immunogenic cell death hypothesis, clinical chemotherapy treatments may result in CD8(+) and CD4(+) T-cell responses against tumor cells. To discover chemotherapy-associated antigens (CAAs), T cells derived from ovarian cancer (OC) patients (who had been treated with appropriate chemotherapy protocols) were interrogated with proteins isolated from primary OC cells. We screened for immunogenicity using two-dimensional electrophoresis gel-eluted OC proteins. Only the selected immunogenic antigens were molecularly characterized by mass-spectrometry-based analysis. Memory T cells that recognized antigens associated with apoptotic (but not live) OC cells were correlated with prolonged survival in response to chemotherapy, supporting the model of chemotherapy-induced apoptosis as an adjuvant of anti-tumor immunity. The strength of both memory CD4(+) and CD8(+) T cells producing either IFN-γ or IL-17 in response to apoptotic OC antigens was also significantly greater in Responders to chemotherapy than in nonresponders. Immunogenicity of some of these antigens was confirmed using recombinant proteins in an independent set of patients. The T-cell interrogation system represents a strategy of reverse tumor immunology that proposes to identify CAAs, which may then be validated as possible prognostic tumor biomarkers or cancer vaccines.
International Journal of Cancer 04/2014; 135(2). DOI:10.1002/ijc.28515 · 5.09 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Group I CD1 proteins are specialized antigen-presenting molecules that present both microbial and self lipid antigens to CD1-restricted alpha/beta T lymphocytes. The production of high levels of gamma interferon and lysis of infected macrophages by lipid-specific T lymphocytes are believed to play pivotal roles mainly in the defense against mycobacterial infections. We previously demonstrated that Mycobacterium tuberculosis and bacillus Calmette-Guérin (Mycobacterium bovis BCG) induce human monocytes to differentiate into CD1- dendritic cells (DC), which cannot present lipid antigens to specific T cells. Here, we show that in human monocytes mycobacteria trigger phosphorylation of p38 mitogen-activated protein kinase to inhibit CD1 expression in DC derived from infected monocytes. Pretreatment with a specific p38 inhibitor renders monocytes insensitive to mycobacterial subversion and allows them to differentiate into CD1+ DC, which are fully capable of presenting lipid antigens to specific T cells. We also report that one of the pathogen recognition receptors triggered by BCG to activate p38 is complement receptor 3 (CR3), as shown by reduced p38 phosphorylation and partial reestablishment of CD1 membrane expression obtained by CR3 blockade before infection. In conclusion, we propose that p38 signaling is a novel pathway exploited by mycobacteria to affect the expression of CD1 antigen-presenting cells and avoid immune recognition.
Infection and immunity 09/2009; 77(11):4947-52. DOI:10.1128/IAI.00607-09 · 3.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: CD4+CD25+Foxp3+ Tregs suppress autoimmune responses. In addition, they limit T cell responses during chronic infection, thereby minimizing T cell-dependent immunopathology. We sought to investigate how Tregs are regulated in the livers of patients chronically infected with HCV, where they control the balance between an adequate protective immune response and suppression of immunopathology. We found that, despite accumulating and proliferating at sites of infection in the livers of patients chronically infected with HCV, Tregs were relatively less expanded than CD4+CD25+Foxp3- effector T cells. The relative lower expansion of intrahepatic Tregs coincided with their upregulation of programmed death-1 (PD-1). PD-1 expression inversely correlated with both Treg proliferation and clinical markers of immune suppression in vivo. Consistent with the possibility that PD-1 controls Tregs, blockade of the interaction between PD-1 and programmed death-1 ligand 1 (PD-L1) enhanced the in vitro expansion and function of Tregs isolated from the livers of patients chronically infected with HCV. Blockade of the interaction between PD-L1 and B7.1 also improved the proliferation of these cells. Interestingly, both PD-1 and phosphorylated STAT-5 were overexpressed in intrahepatic Tregs in a parallel fashion in steady disease conditions, and in an alternate-fluctuating fashion during the course of severe hepatitis reactivation. Notably, PD-L1 blockade upregulated STAT-5 phosphorylation in Tregs ex vivo. These data suggest that PD-L1 negatively regulates Tregs at sites of chronic inflammation by controlling STAT-5 phosphorylation.
The Journal of clinical investigation 03/2009; 119(3):551-64. DOI:10.1172/JCI36604 · 13.22 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We found that the proteome of apoptotic T cells includes prominent fragments of cellular proteins generated by caspases and that a high proportion of distinct T cell epitopes in these fragments is recognized by CD8+ T cells during HIV infection. The frequencies of effector CD8+ T cells that are specific for apoptosis-dependent epitopes correlate with the frequency of circulating apoptotic CD4+ T cells in HIV-1-infected individuals. We propose that these self-reactive effector CD8+ T cells may contribute to the systemic immune activation during chronic HIV infection. The caspase-dependent cleavage of proteins associated with apoptotic cells has a key role in the induction of self-reactive CD8+ T cell responses, as the caspase-cleaved fragments are efficiently targeted to the processing machinery and are cross-presented by dendritic cells. These findings demonstrate a previously undescribed role for caspases in immunopathology.
Nature medicine 01/2008; 13(12):1431-9. DOI:10.1038/nm1679 · 27.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: JC virus (JCV) causes progressive multifocal leukoencephalopathy (PML), characterized by multiple areas of demyelination and attendant loss of brain function. PML is often associated with immunodepression and it is significantly frequent in AIDS patients. The viral genome is divided into early and late genes, between which lies a non-coding control region (NCCR) that regulates JCV replication and presents a great genetic variability. The NCCR of JCV archetype (CY strain) is divided into six regions: A-F containing binding sites for cell factors involved in viral transcription. Deletions and enhancements of these binding sites characterize JCV variants, which could promote viral gene expression and could be more suitable for the onset or development of PML. Therefore, we evaluated by means of polymerase chain reaction (PCR) the presence of JCV genome in cerebrospinal fluid (CSF) of HIV positive and negative subjects both with PML and after sequencing, we analyzed the viral variants found focusing on Sp1 binding sites (box B and D) and up-TAR sequence (box C). It is known that Sp1 activates JCV early promoter and can contribute in maintaining methylation-free CpG islands in active genes, while up-TAR sequence is important for HIV-1 Tat stimulation of JCV late promoter. Our results showed that in HIV-positive subjects all NCCR structures presented enhancements of up-TAR element, whereas in HIV-negative subjects both Sp1 binding sites were always retained. Therefore, we can support the synergism HIV-1/JCV in CNS and we can hypothesize that both Sp1 binding sites could be important to complete JCV replication cycle in absence of HIV-coinfection.
[Show abstract][Hide abstract] ABSTRACT: In human cancer, a role has been suggested for the human polyomavirus BK, primarily associated with tubulointerstitial nephritis and ureteric stenosis in renal transplant recipients, and with hemorrhagic cystitis in bone marrow transplant (BMT) recipients. After the initial infection, primarily unapparent and without clinical signs, the virus disseminates and establishes a persistent infection in the urinary tract and lymphocytes. There is correlative evidence regarding potential role of polyomavirus BK in cancer. In fact, the BK virus (BKV) DNA (complete genome and/or subgenomic fragments containing the early region) is able to transform embryonic fibroblasts and cells cultured from kidney and brain of hamster, mouse, rat, rabbit, and monkey. Nevertheless, transformation of human cells by BKV is inefficient and often abortive. Evidence supporting a possible role for BKV in human cancer has accumulated slowly in recent years, after the advent of polymerase chain reaction (PCR). BKV is known to commonly establish persistent infections in people and to be excreted in the urine by individuals who are asymptomatic, complicating the evaluation of its potential role in development of human cancer. Therefore, there is no certain proof that human polyomavirus BK directly causes the cancer in humans or acts as a cofactor in the pathogenesis of some types of human cancer.
[Show abstract][Hide abstract] ABSTRACT: Hemorrhagic cystitis (HC) is a well-known complication after allogeneic bone marrow transplant (BMT) and can be related to adenovirus or human polyomavirus BK (BKV) infections. In this study a group of 20 patients after allogeneic BMT has been examined. BMT urine samples were analysed for the presence of Adenovirus and BKV DNAby means of polymerase chain reaction (PCR). 5/20 BMT patients developed HC after BMT. The presence of BKV DNA in urine samples was evident in 3/15 patients without HC and in 5/5 patients with HC. In 2/5 HC-patients the BKV DNA was not found after therapy with Cidofovir and Ribavirin. The search for adenovirus DNA in all samples was negative. The analysis of BKV non-coding control region (NCCR) isolated from urine samples revealed a structure very similar to the archetype in all samples. The RFLP (Restriction Fragment Length Polymorphism assay) showed the presence of BKV subtypes I and IV, with the prevalence of subtype I (4/5). This study supports the hypothesis that HC is mainly related to BKV rather than to adenovirus infection in BMT patients. Moreover, since BKV subtype I was predominant, it is reasonable to hypothesize that a specific BKV subtype could be associated with the development of HC.
International journal of immunopathology and pharmacology 04/2005; 18(2):309-16. DOI:10.1016/S0022-5347(05)00850-5 · 1.62 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The JC virus (JCV) is generally considered the etiological agent of progressive multifocal leukoencephalopathy (PML), a demyelinating brain illness, often associated with immunosuppression and significantly frequent in acquired immunodeficiency syndrome (AIDS) patients. The primary infection by JCV is usually asymptomatic and the virus can remain in a latent status in the kidney. As a consequence of immunological alterations of the host, the virus can show a genetic variability in the noncoding control region (NCCR) due to deletions, duplications, and insertions as compared with the archetype. The NCCR of the archetype strain can be divided into six regions, named boxes A to F. In this study, the authors evaluated the presence of the JCV genome in different biological samples, such as urine, peripheral blood mononuclear cells (PBMCs) and cerebral spinal fluid (CSF) by means of polymerase chain reaction (PCR). After sequencing of the PCR fragments, the NCCR structure of isolated JCV strains was analyzed in order to verify the presence of different viral variants. An analysis of the homology and of the multiple alignment of the obtained sequences in comparison with the archetype strain has been carried out. The results indicated the presence of different rearrangements among the analyzed samples. Whereas in the urine, the NCCR structure always appeared very similar to that of the archetype, in the PBMCs and CSF, the NCCR sequences showed specific and characteristic rearrangements as compared to the archetype. These different rearrangements could be correlated with the emerging of an NCCR organization more suitable for the development of PML.
Journal of NeuroVirology 01/2004; 9(6):603-11. DOI:10.1080/13550280390246507 · 2.60 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The distribution of DNA of BK and JC human polyomaviruses (BKV and JCV) was investigated in samples from autopsies of different organs in 2 groups of patients: Human Immunodeficiency Virus -1 (HIV) positive and negative. Samples from various organs were analysed by a nested polymerase chain reaction (PCR) for the non-coding control and for the VP1 regions of both viruses. The results obtained showed that BKV DNA was present in both males and females with a higher prevalence in HIV-positive subject samples (spleen: 33%; kidney: 44%; brain: 22%, uterine cervix:100%; prostatic urethra: 50%). In prostatic urethra samples of HIV-positive subjects, the JCV DNA was revealed in a low percentage (33%), while it was not found at all in uterine cervix samples of both groups. The varying presence of BK and JC viral DNA in the different organs seems to reflect the different pathogenetic attitude of these viruses. JCV was mainly present in the brain (55%), confirming its typical neurotropism and its etiological role in neurological disorders found in immunodeficient patients. BKV, on the other hand, was mainly present in the kidney (44%) and in genital organs (uterine cervix: 100%; prostatic urethra: 50%) with the latter finding favouring the hypothesis of a possible sexual transmission of BKV. Furthermore, our results confirm the crucial role of the immune system in the persistence of human polyomaviruses in the host.
International journal of immunopathology and pharmacology 09/2003; 16(3):269-76. · 1.62 Impact Factor