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1.
J Exp Med ; 216(12): 2778-2799, 2019 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-31601675

RESUMO

Hemophagocytic lymphohistiocytosis (HLH) is characterized by immune dysregulation due to inadequate restraint of overactivated immune cells and is associated with a variable clinical spectrum having overlap with more common pathophysiologies. HLH is difficult to diagnose and can be part of inflammatory syndromes. Here, we identify a novel hematological/autoinflammatory condition (NOCARH syndrome) in four unrelated patients with superimposable features, including neonatal-onset cytopenia with dyshematopoiesis, autoinflammation, rash, and HLH. Patients shared the same de novo CDC42 mutation (Chr1:22417990C>T, p.R186C) and altered hematopoietic compartment, immune dysregulation, and inflammation. CDC42 mutations had been associated with syndromic neurodevelopmental disorders. In vitro and in vivo assays documented unique effects of p.R186C on CDC42 localization and function, correlating with the distinctiveness of the trait. Emapalumab was critical to the survival of one patient, who underwent successful bone marrow transplantation. Early recognition of the disorder and establishment of treatment followed by bone marrow transplant are important to survival.

3.
Front Immunol ; 10: 316, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31031743

RESUMO

Background: Primary Immunodeficiencies (PIDs) are a heterogeneous group of genetic immune disorders. While some PIDs can manifest with more than one phenotype, signs, and symptoms of various PIDs overlap considerably. Recently, novel defects in immune-related genes and additional variants in previously reported genes responsible for PIDs have been successfully identified by Next Generation Sequencing (NGS), allowing the recognition of a broad spectrum of disorders. Objective: To evaluate the strength and weakness of targeted NGS sequencing using custom-made Ion Torrent and Haloplex (Agilent) panels for diagnostics and research purposes. Methods: Five different panels including known and candidate genes were used to screen 105 patients with distinct PID features divided in three main PID categories: T cell defects, Humoral defects and Other PIDs. The Ion Torrent sequencing platform was used in 73 patients. Among these, 18 selected patients without a molecular diagnosis and 32 additional patients were analyzed by Haloplex enrichment technology. Results: The complementary use of the two custom-made targeted sequencing approaches allowed the identification of causative variants in 28.6% (n = 30) of patients. Twenty-two out of 73 (34.6%) patients were diagnosed by Ion Torrent. In this group 20 were included in the SCID/CID category. Eight out of 50 (16%) patients were diagnosed by Haloplex workflow. Ion Torrent method was highly successful for those cases with well-defined phenotypes for immunological and clinical presentation. The Haloplex approach was able to diagnose 4 SCID/CID patients and 4 additional patients with complex and extended phenotypes, embracing all three PID categories in which this approach was more efficient. Both technologies showed good gene coverage. Conclusions: NGS technology represents a powerful approach in the complex field of rare disorders but its different application should be weighted. A relatively small NGS target panel can be successfully applied for a robust diagnostic suspicion, while when the spectrum of clinical phenotypes overlaps more than one PID an in-depth NGS analysis is required, including also whole exome/genome sequencing to identify the causative gene.

4.
Lancet Haematol ; 6(5): e239-e253, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30981783

RESUMO

BACKGROUND: Wiskott-Aldrich syndrome is a rare, life-threatening, X-linked primary immunodeficiency characterised by microthrombocytopenia, infections, eczema, autoimmunity, and malignant disease. Lentiviral vector-mediated haemopoietic stem/progenitor cell (HSPC) gene therapy is a potentially curative treatment that represents an alternative to allogeneic HSPC transplantation. Here, we report safety and efficacy data from an interim analysis of patients with severe Wiskott-Aldrich syndrome who received lentiviral vector-derived gene therapy. METHODS: We did a non-randomised, open-label, phase 1/2 clinical study in paediatric patients with severe Wiskott-Aldrich syndrome, defined by either WAS gene mutation or absent Wiskott-Aldrich syndrome protein (WASP) expression or a Zhu clinical score of 3 or higher. We included patients who had no HLA-identical sibling donor available or, for children younger than 5 years of age, no suitable 10/10 matched unrelated donor or 6/6 unrelated cord blood donor. After treatment with rituximab and a reduced-intensity conditioning regimen of busulfan and fludarabine, patients received one intravenous infusion of autologous CD34+ cells genetically modified with a lentiviral vector encoding for human WAS cDNA. The primary safety endpoints were safety of the conditioning regimen and safety of lentiviral gene transfer into HSPCs. The primary efficacy endpoints were overall survival, sustained engraftment of genetically corrected HSPCs, expression of vector-derived WASP, improved T-cell function, antigen-specific responses to vaccinations, and improved platelet count and mean platelet volume normalisation. This interim analysis was done when the first six patients treated had completed at least 3 years of follow-up. The planned analyses are presented for the intention-to-treat population. This trial is registered with ClinicalTrials.gov (number NCT01515462) and EudraCT (number 2009-017346-32). FINDINGS: Between April 20, 2010, and Feb 26, 2015, nine patients (all male) were enrolled of whom one was excluded after screening; the age range of the eight treated children was 1·1-12·4 years. At the time of the interim analysis (data cutoff April 29, 2016), median follow-up was 3·6 years (range 0·5-5·6). Overall survival was 100%. Engraftment of genetically corrected HSPCs was successful and sustained in all patients. The fraction of WASP-positive lymphocytes increased from a median of 3·9% (range 1·8-35·6) before gene therapy to 66·7% (55·7-98·6) at 12 months after gene therapy, whereas WASP-positive platelets increased from 19·1% (range 4·1-31·0) to 76·6% (53·1-98·4). Improvement of immune function was shown by normalisation of in-vitro T-cell function and successful discontinuation of immunoglobulin supplementation in seven patients with follow-up longer than 1 year, followed by positive antigen-specific response to vaccination. Severe infections fell from 2·38 (95% CI 1·44-3·72) per patient-year of observation (PYO) in the year before gene therapy to 0·31 (0·04-1·11) per PYO in the second year after gene therapy and 0·17 (0·00-0·93) per PYO in the third year after gene therapy. Before gene therapy, platelet counts were lower than 20 × 109 per L in seven of eight patients. At the last follow-up visit, the platelet count had increased to 20-50 × 109 per L in one patient, 50-100 × 109 per L in five patients, and more than 100 × 109 per L in two patients, which resulted in independence from platelet transfusions and absence of severe bleeding events. 27 serious adverse events in six patients occurred after gene therapy, 23 (85%) of which were infectious (pyrexia [five events in three patients], device-related infections, including one case of sepsis [four events in three patients], and gastroenteritis, including one case due to rotavirus [three events in two patients]); these occurred mainly in the first 6 months of follow-up. No adverse reactions to the investigational drug product and no abnormal clonal proliferation or leukaemia were reported after gene therapy. INTERPRETATION: Data from this study show that gene therapy provides a valuable treatment option for patients with severe Wiskott-Aldrich syndrome, particularly for those who do not have a suitable HSPC donor available. FUNDING: Italian Telethon Foundation, GlaxoSmithKline, and Orchard Therapeutics.


Assuntos
Terapia Genética , Vetores Genéticos/genética , Células-Tronco Hematopoéticas/metabolismo , Lentivirus/genética , Síndrome de Wiskott-Aldrich/genética , Síndrome de Wiskott-Aldrich/terapia , Criança , Pré-Escolar , Feminino , Terapia Genética/métodos , Transplante de Células-Tronco Hematopoéticas/métodos , Humanos , Lactente , Itália , Masculino , Mutação , Linfócitos T/imunologia , Linfócitos T/metabolismo , Condicionamento Pré-Transplante/métodos , Resultado do Tratamento , Síndrome de Wiskott-Aldrich/sangue , Síndrome de Wiskott-Aldrich/diagnóstico , Proteína da Síndrome de Wiskott-Aldrich/genética
5.
Front Immunol ; 10: 130, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30837984

RESUMO

We described for the first time a female patient with the simultaneous presence of two homozygous mutations in MYD88 and CARD9 genes presenting with pyogenic bacterial infections, elevated IgE, and persistent EBV viremia. In addition to defective TLR/IL1R-signaling, we described novel functional alterations into the myeloid compartment. In particular, we demonstrated a defective production of reactive oxygen species exclusively in monocytes upon E. coli stimulation, the inability of immature mono-derived DCs (iDCs) to differentiate into mature DCs (mDCs) and the incapacity of mono-derived macrophages (MDMs) to resolve BCG infection in vitro. Our data do not provide any evidence for digenic inheritance in our patient, but rather for the association of two monogenic disorders. This case illustrates the importance of using next generation sequencing (NGS) to determine the most accurate and early diagnosis in atypical clinical and immunological phenotypes, and with particular concern in consanguineous families. Indeed, besides the increased susceptibility to recurrent invasive pyogenic bacterial infections due to MYD88 deficiency, the identification of CARD9 mutations underline the risk of developing invasive fungal infections emphasizing the careful monitoring for the occurrence of fungal infection and the opportunity of long-term antifungal prophylaxis.


Assuntos
Proteínas Adaptadoras de Sinalização CARD/genética , Células Dendríticas/fisiologia , Infecções por Vírus Epstein-Barr/genética , Herpesvirus Humano 4/fisiologia , Imunoglobulina E/sangue , Macrófagos/imunologia , Mutação/genética , Fator 88 de Diferenciação Mieloide/genética , Viremia/genética , Pré-Escolar , Consanguinidade , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Homozigoto , Humanos , Linhagem
7.
Carbohydr Polym ; 207: 720-728, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30600058

RESUMO

The antibacterial activity of the S-unsubstituted- and S-benzyl-substituted-2-mercapto-benzothiazoles 1-4 has been evaluated after complexation with Methyl-ß-Cyclodextrin (Me-ß-CD) or incorporation in solid dispersions based on Pluronic® F-127 and compared with that of the pure compounds. This with the aim to gain further insights on the possible mechanism(s) involved in the CD-mediated enhancement of antimicrobial effectiveness, a promising methodology to overcome the microbial resistance issue. Together with Differential Scanning Calorimetry, FT-IR spectroscopy and X-ray Powder Diffraction investigations, a molecular modeling study focused on compounds 2 and 4 showed that the S-unsubstituted compound 2/Me-ß-CD complex should be more stable than S-benzyl-substituted 4/Me-ß-CD. Only for 1/Me-ß-CD or, particularly, 2/Me-ß-CD complexes, the antibacterial effectiveness was enhanced in the presence of selected bacterial strains. The results herein presented support the mechanisms focusing on the interactions of the bacterial membrane with CD complexes more than those focusing on the improvement of dissolution properties consequent to CD complexation.


Assuntos
Antibacterianos/farmacologia , Benzotiazóis/farmacologia , beta-Ciclodextrinas/química , Antibacterianos/química , Bacillus subtilis/efeitos dos fármacos , Benzotiazóis/química , Composição de Medicamentos , Enterococcus faecalis/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Modelos Moleculares , Poloxâmero/química , Solubilidade , Staphylococcus aureus/efeitos dos fármacos , Tensoativos/química
8.
J Pediatr Hematol Oncol ; 41(4): e266-e269, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30044346

RESUMO

BACKGROUND: Jagunal homolog 1 (JAGN1) gene was identified as a novel responsible for severe congenital neutropenia. The protein encoded by this gene is required for neutrophil differentiation, survival and function in microbial activity. JAGN1-deficient human neutrophils are characterized by alterations in trafficking within the endoplasmic reticulum and golgi compartments because of ultrastructural defects in endoplasmic reticulum and susceptibility to apoptosis. OBSERVATIONS: We report a patient exhibiting an intermittent neutropenia, for which a next-generation sequencing revealed a homozygous mutation in the JAGN1 gene. CONCLUSIONS: The patient extends the clinical variability associated to JAGN1 mutations, and this case highlights the importance of genetic investigations in patients with suspected neutropenia.


Assuntos
Proteínas de Membrana/genética , Neutropenia/congênito , Neutropenia/genética , Pré-Escolar , Sequenciamento de Nucleotídeos em Larga Escala , Homozigoto , Humanos , Masculino , Mutação
9.
Blood ; 132(22): 2362-2374, 2018 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-30254128

RESUMO

ARPC1B is a key factor for the assembly and maintenance of the ARP2/3 complex that is involved in actin branching from an existing filament. Germline biallelic mutations in ARPC1B have been recently described in 6 patients with clinical features of combined immunodeficiency (CID), whose neutrophils and platelets but not T lymphocytes were studied. We hypothesized that ARPC1B deficiency may also lead to cytoskeleton and functional defects in T cells. We have identified biallelic mutations in ARPC1B in 6 unrelated patients with early onset disease characterized by severe infections, autoimmune manifestations, and thrombocytopenia. Immunological features included T-cell lymphopenia, low numbers of naïve T cells, and hyper-immunoglobulin E. Alteration in ARPC1B protein structure led to absent/low expression by flow cytometry and confocal microscopy. This molecular defect was associated with the inability of patient-derived T cells to extend an actin-rich lamellipodia upon T-cell receptor (TCR) stimulation and to assemble an immunological synapse. ARPC1B-deficient T cells additionally displayed impaired TCR-mediated proliferation and SDF1-α-directed migration. Gene transfer of ARPC1B in patients' T cells using a lentiviral vector restored both ARPC1B expression and T-cell proliferation in vitro. In 2 of the patients, in vivo somatic reversion restored ARPC1B expression in a fraction of lymphocytes and was associated with a skewed TCR repertoire. In 1 revertant patient, memory CD8+ T cells expressing normal levels of ARPC1B displayed improved T-cell migration. Inherited ARPC1B deficiency therefore alters T-cell cytoskeletal dynamics and functions, contributing to the clinical features of CID.


Assuntos
Complexo 2-3 de Proteínas Relacionadas à Actina/genética , Mutação em Linhagem Germinativa , Síndromes de Imunodeficiência/genética , Linfócitos T/patologia , Complexo 2-3 de Proteínas Relacionadas à Actina/química , Feminino , Homozigoto , Humanos , Síndromes de Imunodeficiência/patologia , Masculino , Modelos Moleculares , Linhagem , Conformação Proteica , Imunodeficiência Combinada Severa/genética , Imunodeficiência Combinada Severa/patologia , Linfócitos T/metabolismo
10.
Front Immunol ; 9: 113, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29456531

RESUMO

Adenosine deaminase-deficient severe combined immunodeficiency disease (ADA-SCID) is a primary immune deficiency characterized by mutations in the ADA gene resulting in accumulation of toxic compounds affecting multiple districts. Hematopoietic stem cell transplantation (HSCT) from a matched donor and hematopoietic stem cell gene therapy are the preferred options for definitive treatment. Enzyme replacement therapy (ERT) is used to manage the disease in the short term, while a decreased efficacy is reported in the medium-long term. To date, eight cases of lymphomas have been described in ADA-SCID patients. Here we report the first case of plasmablastic lymphoma occurring in a young adult with ADA-SCID on long-term ERT, which turned out to be Epstein-Barr virus associated. The patient previously received infusions of genetically modified T cells. A cumulative analysis of the eight published cases of lymphoma from 1992 to date, and the case here described, reveals a high mortality (89%). The most common form is diffuse large B-cell lymphoma, which predominantly occurs in extra nodal sites. Seven cases occurred in patients on ERT and two after haploidentical HSCT. The significant incidence of immunodeficiency-associated lymphoproliferative disorders and poor survival of patients developing this complication highlight the priority in finding a prompt curative treatment for ADA-SCID.


Assuntos
Adenosina Desaminase/deficiência , Agamaglobulinemia/complicações , Infecções por Vírus Epstein-Barr/complicações , Linfoma Plasmablástico/etiologia , Imunodeficiência Combinada Severa/complicações , Adenosina Desaminase/uso terapêutico , Adolescente , Agamaglobulinemia/tratamento farmacológico , Agamaglobulinemia/etiologia , Terapia de Reposição de Enzimas , Evolução Fatal , Feminino , Humanos , Linfoma Plasmablástico/diagnóstico , Imunodeficiência Combinada Severa/tratamento farmacológico , Imunodeficiência Combinada Severa/etiologia
11.
Front Immunol ; 9: 2767, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30692987

RESUMO

Adenosine deaminase 2 (ADA2) deficiency is an auto-inflammatory disease due to mutations in cat eye syndrome chromosome region candidate 1 (CECR1) gene, currently named ADA2. The disease has a wide clinical spectrum encompassing early-onset vasculopathy (targeting skin, gut and central nervous system), recurrent fever, immunodeficiency and bone marrow dysfunction. Different therapeutic options have been proposed in literature, but only steroids and anti-cytokine monoclonal antibodies (such as tumor necrosis factor inhibitor) proved to be effective. If a suitable donor is available, hematopoietic stem cell transplantation (HSCT) could be curative. Here we describe a case of ADA2 deficiency in a 4-year-old Caucasian girl. The patient was initially classified as autoimmune neutropenia and then she evolved toward an autoimmune lymphoproliferative syndrome (ALPS)-like phenotype. The diagnosis of ALPS became uncertain due to atypical clinical features and normal FAS-induced apoptosis test. She was treated with G-CSF first and subsequently with immunosuppressive drugs without improvement. Only HSCT from a 9/10 HLA-matched unrelated donor, following myeloablative conditioning, completely solved the clinical signs related to ADA2 deficiency. Early diagnosis in cases presenting with hematological manifestations, rather than classical vasculopathy, allows the patients to promptly undergo HSCT and avoid more severe evolution. Finally, in similar cases highly suspicious for genetic disease, it is desirable to obtain molecular diagnosis before performing HSCT, since it can influence the transplant procedure. However, if HSCT has to be performed without delay for clinical indication, related donors should be excluded to avoid the risk of relapse or partial benefit due to a hereditary genetic defect.


Assuntos
Adenosina Desaminase/deficiência , Síndrome Linfoproliferativa Autoimune/terapia , Transplante de Células-Tronco Hematopoéticas , Peptídeos e Proteínas de Sinalização Intercelular/deficiência , Condicionamento Pré-Transplante , Doadores não Relacionados , Adenosina Desaminase/imunologia , Apoptose/efeitos dos fármacos , Apoptose/imunologia , Síndrome Linfoproliferativa Autoimune/enzimologia , Síndrome Linfoproliferativa Autoimune/imunologia , Síndrome Linfoproliferativa Autoimune/patologia , Pré-Escolar , Feminino , Fator Estimulador de Colônias de Granulócitos/administração & dosagem , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Neutropenia/enzimologia , Neutropenia/imunologia , Neutropenia/patologia , Neutropenia/terapia , Transplante Homólogo , Receptor fas/imunologia
12.
Sci Rep ; 7(1): 8153, 2017 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-28811575

RESUMO

Mesenchymal stromal cells (MSCs) represent a key component of bone marrow (BM) microenvironment and display immune-regulatory properties. We performed a detailed analysis of biological/functional properties of BM-MSCs derived from 33 pediatric patients affected by primary immune-deficiencies (PID-MSCs): 7 Chronic Granulomatous Disease (CGD), 15 Wiskott-Aldrich Syndrome (WAS), 11 Severe Combined Immunodeficiency (SCID). Results were compared with MSCs from 15 age-matched pediatric healthy-donors (HD-MSCs). Clonogenic and proliferative capacity, differentiation ability, immunophenotype, immunomodulatory properties were analyzed. WB and RT-qPCR for CYBB, WAS and ADA genes were performed. All PID-MSCs displayed clonogenic and proliferative capacity, morphology and immunophenotype comparable with HD-MSCs. PID-MSCs maintained the inhibitory effect on T- and B-lymphocyte proliferation, except for decreased inhibitory ability of SCID-MSCs at MSC:PBMC ratio 1:10. While HD- and CGD-MSCs were able to inhibit monocyte maturation into immature dendritic cells, in SCID- and WAS-MSCs this ability was reduced. After Toll-like Receptor priming, PID-MSCs displayed in vitro an altered gene expression profile of pro- and anti-inflammatory soluble factors. PID-MSCs displayed lower PPARγ levels and WAS- and SCID-MSCs higher levels of key osteogenic markers, as compared with HD-MSCs. Our results indicate that PID-MSCs may be defective in some functional abilities; whether these defects contribute to disease pathophysiology deserves further investigation.


Assuntos
Síndromes de Imunodeficiência/etiologia , Síndromes de Imunodeficiência/metabolismo , Células-Tronco Mesenquimais/metabolismo , Adipócitos/citologia , Adipócitos/metabolismo , Adolescente , Linfócitos B/imunologia , Linfócitos B/metabolismo , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Diferenciação Celular , Criança , Pré-Escolar , Feminino , Humanos , Imunidade Inata , Lactente , Ativação Linfocitária , Masculino , Células-Tronco Mesenquimais/imunologia , Monócitos/imunologia , Monócitos/metabolismo , Osteoblastos/citologia , Osteoblastos/metabolismo , Células-Tronco , Linfócitos T/imunologia , Linfócitos T/metabolismo , Receptor 3 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo
13.
Curr Med Chem ; 24(34): 3813-3827, 2017 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-28820069

RESUMO

BACKGROUND: Thromboembolic events, principally stroke, represent one of the leading causes of morbidity and mortality among subjects with atrial fibrillation. Chronic kidney disease determines a further increase of thromboembolic events, bleeding and mortality and complicates the pharmacological management of patients with atrial fibrillation, mainly due to the side effects of antiarrhythmic and anticoagulant drugs with renal excretion. Apixaban is a new oral anticoagulant characterized by good bioavailability and renal elimination accounting for only 25%, showing a safety profile and effectiveness in patients with renal impairment. OBJECTIVE: In this manuscript, we reviewed literature data on the use of apixaban in the management of non-valvular atrial fibrillation in patients with renal failure, in order to clarify an often-debated topic in clinical practice. METHOD: A PubMed search was performed on the terms atrial fibrillation, apixaban and renal failure with the aim of identifying relevant manuscripts, large randomized clinical trials, meta-analyses, and current guidelines. RESULTS: Literature data show that apixaban could represent an interesting alternative to warfarin and other selective antagonists of coagulation factors in patients with impaired renal function. About the risk of major bleeding, apixaban appears to be safer than warfarin in the presence of any degree of renal failure. CONCLUSION: Apixaban show to be an effective anticoagulant in patients with atrial fibrillation, even superior to warfarin in reducing the risk of stroke and systemic embolism regardless of the presence of renal insufficiency. Moreover, Food and Drug Administration allows the use of apixaban in patients with end stage renal disease on hemodialysis.


Assuntos
Anticoagulantes/uso terapêutico , Fibrilação Atrial/tratamento farmacológico , Pirazóis/uso terapêutico , Piridonas/uso terapêutico , Insuficiência Renal/complicações , Tromboembolia/prevenção & controle , Anticoagulantes/química , Anticoagulantes/farmacocinética , Fibrilação Atrial/complicações , Testes de Coagulação Sanguínea , Taxa de Filtração Glomerular , Meia-Vida , Humanos , Pirazóis/química , Pirazóis/farmacocinética , Piridonas/química , Piridonas/farmacocinética , Insuficiência Renal/patologia , Índice de Gravidade de Doença , Tromboembolia/etiologia
14.
Clin Immunol ; 178: 20-28, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-26732860

RESUMO

Activated PI3-kinase delta syndrome (APDS) was recently reported as a novel primary immunodeficiency caused by heterozygous gain-of-function mutations in PIK3CD gene. Here we describe immunological studies in a 19year old APDS patient for whom genetic diagnosis was discovered by Whole Exome Sequencing (WES) analysis. In addition to the progressive lymphopenia and defective antibody production we showed that the ability of the patient's B cells to differentiate in vitro is severely reduced. An in depth analysis of the myeloid compartment showed an increased expression of CD83 activation marker on monocytes and mono-derived DC cells. Moreover, monocytes-derived macrophages (MDMs) failed to solve the Mycobacterium bovis bacillus Calmette Guèrin (BCG) infection in vitro. Selective p110δ inhibitor IC87114 restored the MDM capacity to kill BCG in vitro. Our data show that the constitutive activation of Akt-mTOR pathway induces important alterations also in the myeloid compartment providing new insights in order to improve the therapeutic approach in these patients.


Assuntos
Linfócitos B/imunologia , Diferenciação Celular/imunologia , Células Dendríticas/imunologia , Síndromes de Imunodeficiência/imunologia , Macrófagos/imunologia , Adenina/análogos & derivados , Adenina/farmacologia , Diferenciação Celular/genética , Classe I de Fosfatidilinositol 3-Quinases/genética , Classe I de Fosfatidilinositol 3-Quinases/imunologia , Humanos , Síndromes de Imunodeficiência/genética , Técnicas In Vitro , Inflamação , Linfopenia/genética , Linfopenia/imunologia , Macrófagos/efeitos dos fármacos , Masculino , Mycobacterium bovis/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Quinazolinas/farmacologia , Transdução de Sinais , Serina-Treonina Quinases TOR/imunologia , Adulto Jovem
16.
Blood ; 128(1): 45-54, 2016 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-27129325

RESUMO

Adenosine deaminase (ADA) deficiency is a rare, autosomal-recessive systemic metabolic disease characterized by severe combined immunodeficiency (SCID). The treatment of choice for ADA-deficient SCID (ADA-SCID) is hematopoietic stem cell transplant from an HLA-matched sibling donor, although <25% of patients have such a donor available. Enzyme replacement therapy (ERT) partially and temporarily relieves immunodeficiency. We investigated the medium-term outcome of gene therapy (GT) in 18 patients with ADA-SCID for whom an HLA-identical family donor was not available; most were not responding well to ERT. Patients were treated with an autologous CD34(+)-enriched cell fraction that contained CD34(+) cells transduced with a retroviral vector encoding the human ADA complementary DNA sequence (GSK2696273) as part of single-arm, open-label studies or compassionate use programs. Overall survival was 100% over 2.3 to 13.4 years (median, 6.9 years). Gene-modified cells were stably present in multiple lineages throughout follow up. GT resulted in a sustained reduction in the severe infection rate from 1.17 events per person-year to 0.17 events per person-year (n = 17, patient 1 data not available). Immune reconstitution was demonstrated by normalization of T-cell subsets (CD3(+), CD4(+), and CD8(+)), evidence of thymopoiesis, and sustained T-cell proliferative capacity. B-cell function was evidenced by immunoglobulin production, decreased intravenous immunoglobulin use, and antibody response after vaccination. All 18 patients reported infections as adverse events; infections of respiratory and gastrointestinal tracts were reported most frequently. No events indicative of leukemic transformation were reported. Trial details were registered at www.clinicaltrials.gov as #NCT00598481.


Assuntos
Adenosina Desaminase/deficiência , Agamaglobulinemia/terapia , Terapia Genética , Recuperação de Função Fisiológica , Retroviridae , Imunodeficiência Combinada Severa/terapia , Adenosina Desaminase/genética , Adenosina Desaminase/imunologia , Agamaglobulinemia/genética , Agamaglobulinemia/imunologia , Agamaglobulinemia/mortalidade , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Criança , Pré-Escolar , Intervalo Livre de Doença , Feminino , Humanos , Lactente , Masculino , Imunodeficiência Combinada Severa/genética , Imunodeficiência Combinada Severa/imunologia , Imunodeficiência Combinada Severa/mortalidade , Taxa de Sobrevida
18.
J Allergy Clin Immunol ; 136(3): 692-702.e2, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25792466

RESUMO

BACKGROUND: Wiskott-Aldrich syndrome (WAS) is a severe X-linked immunodeficiency characterized by microthrombocytopenia, eczema, recurrent infections, and susceptibility to autoimmunity and lymphomas. Hematopoietic stem cell transplantation is the treatment of choice; however, administration of WAS gene-corrected autologous hematopoietic stem cells has been demonstrated as a feasible alternative therapeutic approach. OBJECTIVE: Because B-cell homeostasis is perturbed in patients with WAS and restoration of immune competence is one of the main therapeutic goals, we have evaluated reconstitution of the B-cell compartment in 4 patients who received autologous hematopoietic stem cells transduced with lentiviral vector after a reduced-intensity conditioning regimen combined with anti-CD20 administration. METHODS: We evaluated B-cell counts, B-cell subset distribution, B cell-activating factor and immunoglobulin levels, and autoantibody production before and after gene therapy (GT). WAS gene transfer in B cells was assessed by measuring vector copy numbers and expression of Wiskott-Aldrich syndrome protein. RESULTS: After lentiviral vector-mediated GT, the number of transduced B cells progressively increased in the peripheral blood of all patients. Lentiviral vector-transduced progenitor cells were able to repopulate the B-cell compartment with a normal distribution of B-cell subsets both in bone marrow and the periphery, showing a WAS protein expression profile similar to that of healthy donors. In addition, after GT, we observed a normalized frequency of autoimmune-associated CD19(+)CD21(-)CD35(-) and CD21(low) B cells and a reduction in B cell-activating factor levels. Immunoglobulin serum levels and autoantibody production improved in all treated patients. CONCLUSIONS: We provide evidence that lentiviral vector-mediated GT induces transgene expression in the B-cell compartment, resulting in ameliorated B-cell development and functionality and contributing to immunologic improvement in patients with WAS.


Assuntos
Subpopulações de Linfócitos B/metabolismo , Terapia Genética/métodos , Transplante de Células-Tronco Hematopoéticas , Proteína da Síndrome de Wiskott-Aldrich/genética , Síndrome de Wiskott-Aldrich/terapia , Autoanticorpos/biossíntese , Fator Ativador de Células B/genética , Fator Ativador de Células B/metabolismo , Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/patologia , Medula Óssea/imunologia , Medula Óssea/metabolismo , Medula Óssea/patologia , Criança , Pré-Escolar , Expressão Gênica , Perfilação da Expressão Gênica , Vetores Genéticos , Células-Tronco Hematopoéticas/imunologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Imunoglobulinas/biossíntese , Imunofenotipagem , Lactente , Lentivirus/genética , Masculino , Proteínas Recombinantes de Fusão/uso terapêutico , Transdução Genética , Condicionamento Pré-Transplante , Transplante Autólogo , Síndrome de Wiskott-Aldrich/genética , Síndrome de Wiskott-Aldrich/imunologia , Síndrome de Wiskott-Aldrich/patologia , Proteína da Síndrome de Wiskott-Aldrich/imunologia
19.
Anticancer Agents Med Chem ; 14(10): 1397-1401, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25173557

RESUMO

Hyperforin is a natural phloroglucinol that has been know for the treatment of depression. Hyperforin displays also antibacterial, antiproliferant and antiangiogenic activity. Synthetic derivatives of hyperforin have also recently been reported to possess increased bioactivity. The clinical applications are limited by the hydrophobic characteristics and the instability of the molecule. In this review we discuss about some of the derivatives of hyperforin (aristoforin, tetrahydrohyperforin and octahydrohyperforin) that demonstrated promising antitumor activity. Among these, octahydrohyperforin also possesses antibacterial activity against both the planktonic and biofilm states of bacteria.


Assuntos
Antibacterianos/farmacologia , Antineoplásicos/farmacologia , Floroglucinol/análogos & derivados , Terpenos/farmacologia , Animais , Linhagem Celular Tumoral , Humanos , Hypericum/química , Floroglucinol/farmacologia , Fármacos Fotossensibilizantes/farmacologia
20.
J Allergy Clin Immunol ; 133(3): 799-806.e10, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24506932

RESUMO

BACKGROUND: Adenosine deaminase (ADA) deficiency causes severe cellular and humoral immune defects and dysregulation because of metabolic toxicity. Alterations in B-cell development and function have been poorly studied. Enzyme replacement therapy (ERT) and hematopoietic stem cell (HSC) gene therapy (GT) are therapeutic options for patients lacking a suitable bone marrow (BM) transplant donor. OBJECTIVE: We sought to study alterations in B-cell development in ADA-deficient patients and investigate the ability of ERT and HSC-GT to restore normal B-cell differentiation and function. METHODS: Flow cytometry was used to characterize B-cell development in BM and the periphery. The percentage of gene-corrected B cells was measured by using quantitative PCR. B cells were assessed for their capacity to proliferate and release IgM after stimulation. RESULTS: Despite the severe peripheral B-cell lymphopenia, patients with ADA-deficient severe combined immunodeficiency showed a partial block in central BM development. Treatment with ERT or HSC-GT reverted most BM alterations, but ERT led to immature B-cell expansion. In the periphery transitional B cells accumulated under ERT, and the defect in maturation persisted long-term. HSC-GT led to a progressive improvement in B-cell numbers and development, along with increased levels of gene correction. The strongest selective advantage for ADA-transduced cells occurred at the transition from immature to naive cells. B-cell proliferative responses and differentiation to immunoglobulin secreting IgM after B-cell receptor and Toll-like receptor triggering were severely impaired after ERT and improved significantly after HSC-GT. CONCLUSIONS: ADA-deficient patients show specific defects in B-cell development and functions that are differently corrected after ERT and HSC-GT.


Assuntos
Adenosina Desaminase/deficiência , Linfócitos B/fisiologia , Terapia de Reposição de Enzimas , Terapia Genética , Transplante de Células-Tronco Hematopoéticas , Adenosina Desaminase/genética , Adenosina Desaminase/uso terapêutico , Adolescente , Fator Ativador de Células B/fisiologia , Linfócitos B/imunologia , Criança , Pré-Escolar , Humanos , Lactente
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