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1.
J Transl Med ; 19(1): 139, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33794925

RESUMO

BACKGROUND: Low T3 syndrome is frequent in patients admitted to intensive care units for critical illness and pneumonia. It has been reported also in patients with COVID-19, Hodgkin disease and chronic lymphocytic leukemia. We analyzed the clinical relevance of Low T3 syndrome in COVID-19 patients and, in particular, in those with associated hematological malignancies. METHODS: Sixty-two consecutive patients, hospitalized during the first wave of SARS-CoV-2 outbreak in Sant'Andrea University Hospital in Rome, were subdivided in 38 patients (Group A), showing low levels of FT3, and in 24 patients (Group B), with normal FT3 serum values. During the acute phase of the disease, we measured serum, radiologic and clinical disease severity markers and scores, in search of possible correlations with FT3 serum values. In addition, in 6 COVID-19 patients, 4 with Low T3 syndrome, including 2 with a hematological malignancy, and 2 with normal FT3 values, we performed, high-dimensional single-cell analysis by mass cytometry, multiplex cytokine assay and gene expression profiling in peripheral blood mononuclear cells (PBMC). RESULTS: Low FT3 serum values were correlated with increased Absolute Neutrophil Count, NLR and dNLR ratios and with reduced total count of CD3+, CD4+ and CD8+ T cells. Low FT3 values correlated also with increased levels of inflammation, tissue damage and coagulation serum markers as well as with SOFA, LIPI and TSS scores. The CyTOF analysis demonstrated reduction of the effector memory and terminal effector subtypes of the CD4+ T lymphocytes. Multiplex cytokine assay indicates that mainly IL-6, IP-10 and MCAF changes are associated with FT3 serum levels, particularly in patients with coexistent hematological malignancies. Gene expression analysis using Nanostring identified four genes differently expressed involved in host immune response, namely CD38, CD79B, IFIT3 and NLRP3. CONCLUSIONS: Our study demonstrates that low FT3 serum levels are associated with severe COVID-19. Our multi-omics approach suggests that T3 is involved in the immune response in COVID-19 and coexistent hematological malignancy and new possible T3 target genes in these patients have been identified.


Assuntos
COVID-19/complicações , Síndromes do Eutireóideo Doente/complicações , Neoplasias Hematológicas , Idoso , Idoso de 80 Anos ou mais , Feminino , Neoplasias Hematológicas/complicações , Neoplasias Hematológicas/genética , Humanos , Itália , Leucócitos Mononucleares , Masculino , Pessoa de Meia-Idade , Análise de Célula Única , Tri-Iodotironina/sangue
2.
J Pathol ; 245(2): 197-208, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29533466

RESUMO

Diabetes is an established risk factor for pancreatic cancer (PaC), together with obesity, a Western diet, and tobacco smoking. The common mechanistic link might be the accumulation of advanced glycation end-products (AGEs), which characterizes all of the above disease conditions and unhealthy habits. Surprisingly, however, the role of AGEs in PaC has not been examined yet, despite the evidence of a tumour-promoting role of receptor for advanced glycation end-products (RAGE), the receptor for AGEs. Here, we tested the hypothesis that AGEs promote PaC through RAGE activation. To this end, we investigated the effects of the AGE Nϵ -carboxymethyllysine (CML) in human pancreatic ductal adenocarcinoma (PDA) cell lines and in a mouse model of Kras-driven PaC interbred with a bioluminescent model of proliferation. Tumour growth was monitored in vivo by bioluminescence imaging and confirmed by histology. CML promoted PDA cell growth and RAGE expression, in a concentration-dependent and time-dependent manner, and activated downstream tumourigenic signalling pathways. These effects were counteracted by RAGE antagonist peptide (RAP). Exogenous AGE administration to PaC-prone mice induced RAGE upregulation in pancreatic intraepithelial neoplasias (PanINs) and markedly accelerated progression to invasive PaC. At 11 weeks of age (6 weeks of CML treatment), PaC was observed in eight of 11 (72.7%) CML-treated versus one of 11 (9.1%) vehicle-treated [control (Ctr)] mice. RAP delayed PanIN development in Ctr mice but failed to prevent PaC promotion in CML-treated mice, probably because of competition with soluble RAGE for binding to AGEs and/or compensatory upregulation of the RAGE homologue CD166/ activated leukocyte cell adhesion molecule, which also favoured tumour spread. These findings indicate that AGEs modulate the development and progression of PaC through receptor-mediated mechanisms, and might be responsible for the additional risk conferred by diabetes and other conditions characterized by increased AGE accumulation. Finally, our data suggest that an AGE reduction strategy, instead of RAGE inhibition, might be suitable for the risk management and prevention of PaC. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Assuntos
Carcinoma in Situ/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Proliferação de Células , Lisina/análogos & derivados , Neoplasias Pancreáticas/metabolismo , Animais , Antineoplásicos/farmacologia , Carcinoma in Situ/tratamento farmacológico , Carcinoma in Situ/genética , Carcinoma in Situ/patologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Complicações do Diabetes/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Genes ras , Humanos , Lisina/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Peptídeos/farmacologia , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Fatores de Risco , Transdução de Sinais , Fatores de Tempo , Carga Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Front Immunol ; 14: 1221587, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38343436

RESUMO

Background: Few data are available about the durability of the response, the induction of neutralizing antibodies, and the cellular response upon the third dose of the anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine in hemato-oncological patients. Objective: To investigate the antibody and cellular response to the BNT162b2 vaccine in patients with hematological malignancy. Methods: We measured SARS-CoV-2 anti-spike antibodies, anti-Omicron neutralizing antibodies, and T-cell responses 1 month after the third dose of vaccine in 93 fragile patients with hematological malignancy (FHM), 51 fragile not oncological subjects (FNO) aged 80-92, and 47 employees of the hospital (healthcare workers, (HW), aged 23-66 years. Blood samples were collected at day 0 (T0), 21 (T1), 35 (T2), 84 (T3), 168 (T4), 351 (T pre-3D), and 381 (T post-3D) after the first dose of vaccine. Serum IgG antibodies against S1/S2 antigens of SARS-CoV-2 spike protein were measured at every time point. Neutralizing antibodies were measured at T2, T3 (anti-Alpha), T4 (anti-Delta), and T post-3D (anti-Omicron). T cell response was assessed at T post-3D. Results: An increase in anti-S1/S2 antigen antibodies compared to T0 was observed in the three groups at T post-3D. After the third vaccine dose, the median antibody level of FHM subjects was higher than after the second dose and above the putative protection threshold, although lower than in the other groups. The neutralizing activity of antibodies against the Omicron variant of the virus was tested at T2 and T post-3D. 42.3% of FHM, 80,0% of FNO, and 90,0% of HW had anti-Omicron neutralizing antibodies at T post-3D. To get more insight into the breadth of antibody responses, we analyzed neutralizing capacity against BA.4/BA.5, BF.7, BQ.1, XBB.1.5 since also for the Omicron variants, different mutations have been reported especially for the spike protein. The memory T-cell response was lower in FHM than in FNO and HW cohorts. Data on breakthrough infections and deaths suggested that the positivity threshold of the test is protective after the third dose of the vaccine in all cohorts. Conclusion: FHM have a relevant response to the BNT162b2 vaccine, with increasing antibody levels after the third dose coupled with, although low, a T-cell response. FHM need repeated vaccine doses to attain a protective immunological response.


Assuntos
COVID-19 , Neoplasias Hematológicas , Glicoproteína da Espícula de Coronavírus , Humanos , Vacinas contra COVID-19 , Vacina BNT162 , COVID-19/prevenção & controle , SARS-CoV-2 , Anticorpos Neutralizantes , Anticorpos Antivirais
5.
Sci Rep ; 11(1): 671, 2021 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-33436662

RESUMO

We developed a novel reporter transgenic zebrafish model called MITO-Luc/GFP zebrafish in which GFP and luciferase expression are under the control of the master regulator of proliferation NF-Y. In MITO-Luc/GFP zebrafish it is possible to visualize cell proliferation in vivo by fluorescence and bioluminescence. In this animal model, GFP and luciferase expression occur in early living embryos, becoming tissue specific in juvenile and adult zebrafish. By in vitro and ex vivo experiments we demonstrate that luciferase activity in adult animals occurs in intestine, kidney and gonads, where detectable proliferating cells are located. Further, by time lapse experiments in live embryos, we observed a wave of GFP positive cells following fin clip. In adult zebrafish, in addition to a bright bioluminescence signal on the regenerating tail, an early unexpected signal coming from the kidney occurs indicating not only a fin cell proliferation, but also a systemic response to tissue damage. Finally, we observed that luciferase activity was inhibited by anti-proliferative interventions, i.e. 5FU, cell cycle inhibitors and X-Rays. In conclusion, MITO-Luc/GFP zebrafish is a novel animal model that may be crucial to assess the spatial and temporal evolution of cell proliferation in vivo.


Assuntos
Animais Geneticamente Modificados/crescimento & desenvolvimento , Proliferação de Células , Evolução Molecular , Proteínas de Fluorescência Verde/metabolismo , Luciferases/metabolismo , Análise Espaço-Temporal , Peixe-Zebra/crescimento & desenvolvimento , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/metabolismo , Proteínas de Fluorescência Verde/genética , Luciferases/genética , Regeneração , Peixe-Zebra/genética , Peixe-Zebra/metabolismo
6.
Mol Oncol ; 15(2): 579-595, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33159833

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer. Most patients present with advanced disease at diagnosis, which only permits palliative chemotherapeutic treatments. RNA dysregulation is a hallmark of most human cancers, including PDAC. To test the impact of RNA processing dysregulation on PDAC pathology, we performed a bioinformatics analysis to identify RNA-binding proteins (RBPs) associated with prognosis. Among the 12 RBPs associated with progression-free survival, we focused on MEX3A because it was recently shown to mark an intestinal stem cell population that is refractory to chemotherapeutic treatments, a typical feature of PDAC. Increased expression of MEX3A was correlated with higher disease stage in PDAC patients and with tumor development in a mouse model of PDAC. Depletion of MEX3A in PDAC cells enhanced sensitivity to chemotherapeutic treatment with gemcitabine, whereas its expression was increased in PDAC cells selected upon chronic exposure to the drug. RNA-sequencing analyses highlighted hundreds of genes whose expression is sensitive to MEX3A expression, with significant enrichment in cell cycle genes. MEX3A binds to its target mRNAs, like cyclin-dependent kinase 6 (CDK6), and promotes their stability. Accordingly, knockdown of MEX3A caused a significant reduction in PDAC cell proliferation and in progression to the S phase of the cell cycle. These findings uncover a novel role for MEX3A in the acquisition and maintenance of chemoresistance by PDAC cells, suggesting that it may represent a novel therapeutic target for PDAC.


Assuntos
Carcinoma Ductal Pancreático/metabolismo , Desoxicitidina/análogos & derivados , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/metabolismo , Neoplasias Pancreáticas/metabolismo , Fosfoproteínas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Linhagem Celular Tumoral , Desoxicitidina/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Neoplasias Experimentais/diagnóstico , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/genética , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Fosfoproteínas/genética , Prognóstico , Proteínas de Ligação a RNA/genética , Gencitabina
7.
Cell Death Dis ; 12(11): 1019, 2021 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-34716309

RESUMO

Clinical outcomes of COVID-19 patients are worsened by the presence of co-morbidities, especially cancer leading to elevated mortality rates. SARS-CoV-2 infection is known to alter immune system homeostasis. Whether cancer patients developing COVID-19 present alterations of immune functions which might contribute to worse outcomes have so far been poorly investigated. We conducted a multi-omic analysis of immunological parameters in peripheral blood mononuclear cells (PBMCs) of COVID-19 patients with and without cancer. Healthy donors and SARS-CoV-2-negative cancer patients were also included as controls. At the infection peak, cytokine multiplex analysis of blood samples, cytometry by time of flight (CyTOF) cell population analyses, and Nanostring gene expression using Pancancer array on PBMCs were performed. We found that eight pro-inflammatory factors (IL-6, IL-8, IL-13, IL-1ra, MIP-1a, IP-10) out of 27 analyzed serum cytokines were modulated in COVID-19 patients irrespective of cancer status. Diverse subpopulations of T lymphocytes such as CD8+T, CD4+T central memory, Mucosal-associated invariant T (MAIT), natural killer (NK), and γδ T cells were reduced, while B plasmablasts were expanded in COVID-19 cancer patients. Our findings illustrate a repertoire of aberrant alterations of gene expression in circulating immune cells of COVID-19 cancer patients. A 19-gene expression signature of PBMCs is able to discriminate COVID-19 patients with and without solid cancers. Gene set enrichment analysis highlights an increased gene expression linked to Interferon α, γ, α/ß response and signaling which paired with aberrant cell cycle regulation in cancer patients. Ten out of the 19 genes, validated in a real-world consecutive cohort, were specific of COVID-19 cancer patients independently from different cancer types and stages of the diseases, and useful to stratify patients in a COVID-19 disease severity-manner. We also unveil a transcriptional network involving gene regulators of both inflammation response and proliferation in PBMCs of COVID-19 cancer patients.


Assuntos
Anticorpos Antivirais/sangue , COVID-19/imunologia , Citocinas/sangue , Leucócitos Mononucleares/imunologia , Neoplasias/imunologia , COVID-19/patologia , Estudos de Casos e Controles , Feminino , Humanos , Leucócitos Mononucleares/citologia , Masculino , Neoplasias/patologia
8.
J Exp Clin Cancer Res ; 39(1): 152, 2020 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-32778157

RESUMO

BACKGROUND: Type 1 and 2 diabetes confer an increased risk of pancreatic cancer (PaC) of similar magnitude, suggesting a common mechanism. The recent finding that PaC incidence increases linearly with increasing fasting glucose levels supports a central role for hyperglycaemia, which is known to cause carbonyl stress and advanced glycation end-product (AGE) accumulation through increased glycolytic activity and non-enzymatic reactions. This study investigated the impact of hyperglycaemia on invasive tumour development and the underlying mechanisms involved. METHODS: Pdx1-Cre;LSL-KrasG12D/+ mice were interbred with mitosis luciferase reporter mice, rendered diabetic with streptozotocin and treated or not with carnosinol (FL-926-16), a selective scavenger of reactive carbonyl species (RCS) and, as such, an inhibitor of AGE formation. Mice were monitored for tumour development by in vivo bioluminescence imaging. At the end of the study, pancreatic tissue was collected for histology/immunohistochemistry and molecular analyses. Mechanistic studies were performed in pancreatic ductal adenocarcinoma cell lines challenged with high glucose, glycolysis- and glycoxidation-derived RCS, their protein adducts AGEs and sera from diabetic patients. RESULTS: Cumulative incidence of invasive PaC at 22 weeks of age was 75% in untreated diabetic vs 25% in FL-926-16-gtreated diabetic and 8.3% in non-diabetic mice. FL-926-16 treatment suppressed systemic and pancreatic carbonyl stress, extracellular signal-regulated kinases (ERK) 1/2 activation, and nuclear translocation of Yes-associated protein (YAP) in pancreas. In vitro, RCS scavenging and AGE elimination completely inhibited cell proliferation stimulated by high glucose, and YAP proved essential in mediating the effects of both glucose-derived RCS and their protein adducts AGEs. However, RCS and AGEs induced YAP activity through distinct pathways, causing reduction of Large Tumour Suppressor Kinase 1 and activation of the Epidermal Growth Factor Receptor/ERK signalling pathway, respectively. CONCLUSIONS: An RCS scavenger and AGE inhibitor prevented the accelerating effect of diabetes on PainINs progression to invasive PaC, showing that hyperglycaemia promotes PaC mainly through increased carbonyl stress. In vitro experiments demonstrated that both circulating RCS/AGEs and tumour cell-derived carbonyl stress generated by excess glucose metabolism induce proliferation by YAP activation, hence providing a molecular mechanism underlying the link between diabetes and PaC (and cancer in general).


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Complicações do Diabetes/patologia , Diabetes Mellitus Experimental/complicações , Produtos Finais de Glicação Avançada/metabolismo , Estresse Oxidativo , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Complicações do Diabetes/etiologia , Complicações do Diabetes/metabolismo , Camundongos , Camundongos Transgênicos , Mutação , Neoplasias Pancreáticas/etiologia , Neoplasias Pancreáticas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas de Sinalização YAP
9.
Front Pharmacol ; 10: 235, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30930779

RESUMO

Preclinical animal models are valuable tools to improve treatments of malignant diseases, being an intermediate step of experimentation between cell culture and human clinical trials. Among different animal models frequently used in cancer research are mouse and, more recently, zebrafish models. Indeed, most of the cellular pathways are highly conserved between human, mouse and zebrafish, thus rendering these models very attractive. Recently, several transgenic reporter mice and zebrafishes have been generated in which the luciferase reporter gene are placed under the control of a promoter whose activity is strictly related to specific cancer cellular processes. Other mouse models have been generated by the cDNA luciferase knockin in the locus of a gene whose expression/activity has increased in cancer. Using BioLuminescence Imaging (BLI), we have now the opportunity to spatiotemporal visualize cell behaviors, among which proliferation, apoptosis, migration and immune responses, in any body district in living animal in a time frame process. We provide here a review of the available models to visualized cancer and cancer-associated events in living animals by BLI and as they have been successful in identifying new stages of early tumor progression, new interactions between different tissues and new therapeutic responsiveness.

10.
Ann Anat ; 207: 2-8, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26704357

RESUMO

Transgenic mouse models designed to recapitulate genetic and pathologic aspects of cancer are useful to study early stages of disease as well as its progression. Among several, two of the most sophisticated models for pancreatic ductal adenocarcinoma (PDAC) are the LSL-Kras(G12D/+);Pdx-1-Cre (KC) and LSL-Kras(G12D/+);LSL-Trp53(R172H/+);Pdx-1-Cre (KPC) mice, in which the Cre-recombinase regulated by a pancreas-specific promoter activates the expression of oncogenic Kras alone or in combination with a mutant p53, respectively. Non-invasive in vivo imaging offers a novel approach to preclinical studies introducing the possibility to investigate biological events in the spatio/temporal dimension. We recently developed a mouse model, MITO-Luc, engineered to express the luciferase reporter gene in cells undergoing active proliferation. In this model, proliferation events can be visualized non-invasively by bioluminescence imaging (BLI) in every body district in vivo. Here, we describe the development and characterization of MITO-Luc-KC- and -KPC mice. In these mice we have now the opportunity to follow PDAC evolution in the living animal in a time frame process. Moreover, by relating in vivo and ex vivo BLI and histopathological data we provide evidence that these mice could represents a suitable tool for pancreatic cancer preclinical studies. Our data also suggest that aberrant proliferation events take place early in pancreatic carcinogenesis, before tumour appearance.


Assuntos
Modelos Animais de Doenças , Luciferases/genética , Camundongos Transgênicos/genética , Microscopia de Fluorescência/métodos , Neoplasias Pancreáticas/patologia , Lesões Pré-Cancerosas/patologia , Animais , Carcinogênese/genética , Genes Reporter , Humanos , Medições Luminescentes/métodos , Proteínas Luminescentes/genética , Camundongos Endogâmicos C57BL , Estadiamento de Neoplasias , Neoplasias Pancreáticas/genética , Lesões Pré-Cancerosas/genética
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