Localization of endogenous STX11 & Munc18-2 in CTL

Cytotoxic T-lymphocytes (CTL) kill their targets by cytolytic granule secretion at the immunological synapse. The Sec/Munc protein, Munc18-2, and its binding partner Syntaxin 11 (STX11) are both required for granule secretion, with mutations in either leading to the primary immunodeficiency, Familial Haemophagocytic Lymphohistiocytosis (FHL4 and 5). Understanding how Munc18-2 and STX11 function in CTL has been hampered by not knowing the endogenous localization of these proteins. Using a novel FHL5 Munc18-2 mutation that results in loss of protein, cytotoxicity and degranulation together with CTL from an FHL4 patient lacking STX11, enabled us to localize endogenous STX11 and Munc18-2 in CTL. Munc18-2 localized predominantly to cytolytic granules with low levels associated with the plasma membrane where STX11 localized. Importantly, while Munc18-2 localization is unaffected by the absence of STX11 in FHL4 CTL, STX11 is lost from the plasma membrane in FHL5 CTL lacking Munc18-2. These findings support a role for Munc18-2 in chaperoning STX11 to the plasma membrane where the final fusion events involved in secretion occur.his work was funded by grants from the Wellcome Trust [075880], [100140] and [097024/B/11/Z] (G.M.G); a Daimler Benz scholarship (Y.H.); the Kendal-Dixon Fund (Y.H.); the Cambridge European Trust (Y.H.); King’s College (Cambridge) (Y.H.); the National Institute for Health Research Biomedical Research Center (G.M.G); Antonio Pinzino – Associazione per la Ricerca sulle Sindromi Emofagocitiche (M.A.); Associazione Italiana Ricerca istiocitosi (M.A.); and Italian Ministry of Health, Progetti dioe ricerca finalizzata 2008, Bando Malattie Rare RF-TOS-2008-1219488 (M.A.).This is the final version of the article. It first appeared from Wiley via http://dx.doi.org/10.1111/tra.1233

Effectiveness of Preventive Measures in Keeping Low Prevalence of SARS-CoV-2 Infection in Health Care Workers in a Referral Children’s Hospital in Southern Italy

The coronavirus disease 2019 (COVID-19) pandemic now represents a major threat to public health. Health care workers (HCW) are exposed to biological risk. Little is currently known about the risk of HCW operating in pediatric wards for SARS-CoV-2 infection. The aim is to assess the prevalence of SARS-CoV-2 infection in HCW in a third-level children’s hospital in Southern Italy. An observational cohort study of all asymptomatic HCW (physician, technicians, nurses, and logistic and support operators) was conducted. HCW were screened, on a voluntary basis, for SARS-CoV-2 by RT-PCR on nasopharyngeal swab performed during the first wave of COVID-19. The study was then repeated, with the same modalities, at a 7-month interval, during the “second wave” of the COVID-19 pandemic. At the initial screening between 7 and 24 April 2020, 525 HCW were tested. None of them tested positive. At the repeated screening, conducted between 9 and 20 November 2020, 627 HCW were tested, including 61 additional ones resulting from COVID-emergency recruitment. At this second screening, eight subjects (1.3%) tested positive, thus being diagnosed as asymptomatic carriers of SARS-CoV-2. They were one physician, five nurses, and two HCW from the logistic/support services. They were employed in eight different wards/services. In all cases, the epidemiological investigation showed convincing evidence that the infection was acquired through social contacts. The study revealed a very low circulation of SARS-CoV-2 infection in HCW tested with RT-PCR. All the infections documented in the second wave of epidemic of SARS-CoV-2 were acquired outside of the workplace, confirming that in a pediatric hospital setting, HCW education, correct use of personal protective equipment, and separation of the COVID-patient pathway and staff flow may minimize the risk derived from occupational exposure

Biosimilars in the management of neutropenia: focus on filgrastim

Advances in chemotherapy and surgery allows the majority of patients to survive cancer diseases. Yet, the price may be a proportion of patients dying of complications due to treatment-induced infectious complications, such as neutropenia. With the aim of decreasing morbidity and mortality related to infectious complications, recombinant human granulocyte colony-stimulating factor (G-CSF), filgrastim, and pegylated filgrastim have been used to reduce time and degree of neutropenia. A biosimilar is a copy of an approved original biologic medicine whose data protection has expired. The patent for filgrastim expired in Europe in 2006 and in the US in 2013. This review analyses the available evidence to be considered in order to design a strategy of use of G-CSF and its biosimilars. The clinical and safety outcomes of biosimilars are well within the range of historically reported data for originator filgrastim. This underscores the clinical effectiveness and safety of biosimilar filgrastim in daily clinical practice. Biosimilars can play an important role by offering the opportunity to reduce costs, thus contributing to the financial sustainability of treatment programs

NK-dependent DC maturation is mediated by TNFα and IFNγ released upon engagement of the NKp30 triggering receptor

Abstract Natural killer (NK) cells were recently shown to play a relevant role in the process of dendritic cell (DC) maturation. This function is exerted either by direct DC stimulation or through killing those DCs that did not properly acquire a mature phenotype. While killing of immature DCs is dependent on the function of the NKp30 triggering receptor, the mechanism by which NK cells induce DC maturation is still unclear. In this study, we show that also the NK-mediated induction of DC maturation is dependent on NKp30. Upon NK/DC interaction, resulting in NKp30 engagement, NK cells produced tumor necrosis factor α (TNFα) (and interferon γ [IFNγ]) that, in turn, promoted DC maturation. Masking of NKp30 with specific monoclonal antibodies (mAbs) strongly reduced maturation of DCs cocultured with NK cells. In addition, supernatant from NK cells stimulated via NKp30 induced DC maturation, and this effect was neutralized by anti-TNFα antibodies (Abs). This NKp30 function is controlled by the HLA-specific inhibitory NK receptors. Accordingly, the ability to promote maturation was essentially confined to NK cells expressing the killer immunoglobulin-like receptor–negative (KIR–) NKG2Adull phenotype. Finally, the analysis of perforin-deficient NK cells allowed the dissection of the 2 NKp30-mediated NK-cell functions, since NKp30 could induce cytokine-dependent DC maturation in the absence of NK-mediated DC killing

Limited Additive Diagnostic Impact of Isolated Gastrointestinal Involvement for the Triage of Children with Suspected COVID-19

The strategy for the selection of patients with a suspected SARS-CoV-2 infection is relevant for the organization of a children’s hospital to provide optimal separation into COVID-19 and non-COVID-19 areas and pathways. We analyzed the proportion of children with COVID-19 presenting with gastrointestinal (GI) symptoms in 137 consecutive patients admitted between January 2020 and August 2021. GI symptoms were present as follows: diarrhea in 35 patients (26%), vomiting in 16 (12%), and both of them in five (3%); the combination of fever, respiratory symptoms, and diarrhea was observed in 16 patients (12%). Of the 676 adult patients with COVID-19 admitted to our hospital in the same time interval, 62 (9.2%) had diarrhea, 30 (4.4%) had vomiting, and 11 (1.6%) had nausea; only one patient, a 38-year-old male, presented with isolated GI symptoms at the diagnosis. Although diarrhea was observed in one quarter of cases, one-half of them had the complete triad of fever, respiratory syndrome, and diarrhea, and only five had isolated diarrhea, of which two were diagnosed with a Campylobacter infection. The occurrence of either respiratory symptoms or gastrointestinal symptoms in our patients was not related to the patient age, while younger children were more likely to have a fever. Of the 137 patients, 73 (53%) could be tested for their serum level of SARS-CoV-2 specific IgG antibodies. The observed titer ranged between 0 (n = 3) and 1729 BAU/mL (median, 425 BAU/mL). Of 137 consecutive patients with COVID-19 admitted to our referral children’s hospital, only three presented with an isolated GI manifestation. It is interesting to note that this finding turned out to be fully in keeping with what was observed on adult patients with COVID-19 in our hospital. The additive diagnostic impact of gastrointestinal involvement for the triage of children with suspected COVID-19 appears limited

Acute leukemia in association with Langerhans cell histiocytosis

Langerhans cell histiocytosis (LCH) and malignancy occurring in the same individual is unusual and has generally been the subject of isolated case reports. To better define the occurrence of these events a registry of cases with synchronous or asynchronous LCH and malignancy was developed with the cooperation of the Histiocyte Society. In 1991 the Histiocyte Society surveyed its members requesting information on cases in which LCH was associated with malignancy. The questionnaire was mailed to all members of the society and specifically requested information on the clinical and laboratory features of the cases, disease evolution, and response to therapy. Retrospective reporting was allowed. With this initial data, an ongoing registry of LCH patients with associated malignancy was begun of such cases, including evolution and response to therapy. Twenty-seven patients were enrolled during the first year of the registry, of whom 4 patients had the association of LCH with a malignant lymphoma and 10 cases had an association of LCH with other types of solid tumor. The remaining 13 patients had the association of LCH with acute leukemia. In five cases, LCH was associated with acute lymphoblastic leukemia FAB L1 (ALL). In four cases the ALL preceded the LCH by 6 months to 1 year. In four of five patients the LCH was localized; in two instances the LCH was treated with chemotherapy. In all cases the leukemia was treated according to local standard ALL protocols and in one case autologous bone marrow transplantation (ABMT) was performed at relapse. Three patients are free of leukemia, one of whom has persistent localized LCH of the skin. Two patients died of the ALL, one of whom was free of the LCH at the time of death. In eight instances LCH was reported in association with acute myeloid leukemia (AML). Six of these patients had a generalized form of LCH. In seven the diagnosis of LCH preceded the diagnosis of leukemia by more than 2 years (median 4 years). In the remaining patient both diagnoses were made concurrently. In all seven cases in whom LCH was the initial diagnosis the treatment consisted of chemotherapy and/or radiotherapy. Seven patients died from the AML, five without evidence of LCH. The temporal patterns of the LCH-ALL and LCH-AML associations are distinct with ALL usually preceding the diagnosis of LCH and AML succeeding it. Such a pattern is suggestive that in cases of ALL the LCH may be a reactive process while in cases of AML occurring after LCH the primary LCH therapy may play an inductive role in the leukemia. © 1994 Wiley-Liss, Inc

Deregulation of Ion Channel and Transporter Encoding Genes in Pediatric Gliomas

Brain tumors, including the majority gliomas, are the leading cause of cancer-related death in children. World Health Organization has divided pediatric brain tumors into different grades and, based upon cDNA microarray data identifying gene expression profiles (GEPs), it has become evident in the last decade that the various grades involve different types of genetic alterations. However, it is not known whether ion channel and transporter genes, intimately involved in brain functioning, are associated with such GEPs. We determined the GEPs in an available cohort of 10 pediatric brain tumors initially by comparing the data obtained from four primary tumor samples and corresponding short-term cultures. The correspondence between the two types of samples was statistically significant. We then performed bioinformatic analyses on those samples (a total of nine) which corresponded to tumors of glial origin, either tissues or cell cultures, depending on the best “RNA integrity number.” We used R software to evaluate the genes which were differentially expressed (DE) in gliomas compared with normal brain. Applying a p-value below 0.01 and fold change ≥4, led to identification of 2284 DE genes. Through a Functional Annotation Analysis (FAA) using the NIH-DAVID software, the DE genes turned out to be associated mainly with: immune/inflammatory response, cell proliferation and survival, cell adhesion and motility, neuronal phenotype, and ion transport. We have shown that GEPs of pediatric brain tumors can be studied using either primary tumor samples or short-term cultures with similar results. From FAA, we concluded that, among DE genes, pediatric gliomas show a strong deregulation of genes related to ion channels and transporters

Foreign children with cancer in Italy

<p>Abstract</p> <p>Background</p> <p>There has been a noticeable annual increase in the number of children coming to Italy for medical treatment, just like it has happened in the rest of the European Union. In Italy, the assistance to children suffering from cancer is assured by the current network of 54 centres members of the Italian Association of Paediatric Haematology and Oncology (AIEOP), which has kept records of all demographic and clinical data in the database of Mod.1.01 Registry since 1989.</p> <p>Methods</p> <p>We used the information stored in the already mentioned database to assess the impact of immigration of foreign children with cancer on centres' activity, with the scope of drawing a map of the assistance to these cases.</p> <p>Results</p> <p>Out of 14,738 cases recorded by all centres in the period from 1999 to 2008, 92.2% were born and resident in Italy, 4.1% (608) were born abroad and living abroad and 3.7% (538) were born abroad and living in Italy. Foreign children cases have increased over the years from 2.5% in 1999 to. 8.1% in 2008.</p> <p>Most immigrant children came from Europe (65.7%), whereas patients who came from America, Asia and Oceania amounted to 13.2%, 10.1%, 0.2%, respectively. The immigrant survival rate was lower compared to that of children who were born in Italy. This is especially true for acute lymphoblastic leukaemia patients entered an AIEOP protocol, who showed a 10-years survival rate of 71.0% vs. 80.7% (p < 0.001) for immigrants and patients born in Italy, respectively.</p> <p>Conclusions</p> <p>Children and adolescents are an increasingly important part of the immigration phenomenon, which occurs in many parts of the world. In Italy the vast majority of children affected by malignancies are treated in AIEOP centres. Since immigrant children are predominantly treated in northern Italy, these centres have developed a special expertise in treating immigrant patients, which is certainly very useful for the entire AIEOP network.</p

Genetic predisposition to hemophagocytic lymphohistiocytosis: Report on 500 patients from the Italian registry.

BACKGROUND: Hemophagocytic lymphohistiocytosis (HLH) is a rare life-threatening disease affecting mostly children but also adults and characterized by hyperinflammatory features. A subset of patients, referred to as having familial hemophagocytic lymphohistiocytosis (FHL), have various underlying genetic abnormalities, the frequencies of which have not been systematically determined previously. OBJECTIVE: This work aims to further our understanding of the pathogenic bases of this rare condition based on an analysis of our 25 years of experience. METHODS: From our registry, we have analyzed a total of 500 unselected patients with HLH. RESULTS: Biallelic pathogenic mutations defining FHL were found in 171 (34%) patients; the proportion of FHL was much higher (64%) in patients given a diagnosis during the first year of life. Taken together, mutations of the genes PRF1 (FHL2) and UNC13D (FHL3) accounted for 70% of cases of FHL. Overall, a genetic diagnosis was possible in more than 90% of our patients with FHL. Perforin expression and the extent of degranulation have been more useful for diagnosing FHL than hemophagocytosis and the cytotoxicity assay. Of 281 (56%) patients classified as having "sporadic" HLH, 43 had monoallelic mutations in one of the FHL-defining genes. Given this gene dosage effect, FHL is not strictly recessive. CONCLUSION: We suggest that the clinical syndrome HLH generally results from the combined effects of an exogenous trigger and genetic predisposition. Within this combination, different weights of exogenous and genetic factors account for the wide disease spectrum that ranges from HLH secondary to severe infection to FHL.Supported by grants from Associazione Italiana Ricerca Istiocitosi (AIRI), Associazione Ciemmeesse-Girotondo per il Meyer, Ministero della Salute (Bando Malattie Rare RF-TOS-2008-1219488), and the Seventh Framework Programme (FP7) of the European Commission (“FIGHT-HLH” Project no. 306124 to M.A.). Disclosure of potential conflict of interest: D. Pende receives royalties paid to her institution. G. M. Griffiths has received research support from the Wellcome Trust. L. Luzzatto is on the Alexion Pharmaceuticals SAB and has received consultancy fees from GlaxoSmithKline.This is the final version of the article. It first appeared from Elsevier via http://dx.doi.org/10.1016/j.jaci.2015.06.04