Background and Purpose
Waning immunity and the surge of SARS-CoV-2 variants are responsible for breakthrough infections, i.e., infections in fully vaccinated individuals. Although the majority of ...vaccinated infected subjects report mild or no symptoms, some others require hospitalization. The clinical and immunological features of vaccinated hospitalized COVID-19 patients are currently unknown.
Methods
Twenty-nine unvaccinated and 36 vaccinated hospitalized COVID-19 patients were prospectively enrolled and clinical and laboratory data were gathered. Immunophenotyping of leukocytes’ subsets, T and B cell SARS-CoV-2-specific responses were evaluated via flow cytometry. Anti-IFN-α autoantibodies were measured via ELISA.
Results
Despite vaccinated patients were older and with more comorbidities, unvaccinated subjects showed higher levels of pro-inflammatory markers, more severe disease, and increased mortality rate. Accordingly, they presented significant alterations in the circulating leukocyte composition, typical of severe COVID-19. Vaccinated patients displayed higher levels of anti-Spike IgGs and Spike-specific B cells. Of all participants, survivors showed higher levels of anti-Spike IgGs and Spike-specific CD4+ T cells than non-survivors. At hospital admission, 6 out of 65 patients (9.2%) displayed high serum concentrations of autoantibodies targeting IFN-α. Remarkably, 3 were unvaccinated and eventually died, while the other 3 were vaccinated and survived.
Conclusion
Despite more severe pre-existing clinical conditions, vaccinated patients have good outcome. A rapid activation of anti-SARS-CoV-2-specific immunity is fundamental for the resolution of the infection. Therefore, prior immunization through vaccination provides a significant contribution to prevention of disease worsening and can even overcome the presence of high-risk factors (i.e., older age, comorbidities, anti-IFN-α autoantibodies).
Measurable residual disease (MRD) is a powerful predictor of outcome in acute myeloid leukemia. In the early phases of treatment, MRD refines initial disease risk stratification and is used for the ...allocation to allogeneic transplant. Despite its well-established role, a relatively high fraction of patients eventually relapses albeit achieving MRDneg status. The aim of this work was to assess specifically the influence of baseline features and treatment intensity on the predictive value of an MRDneg status, particularly focusing on MRD2, measured after two consecutive chemotherapy cycles. Among baseline features, younger MRD2neg patients (<55 years) had a significantly longer disease-free survival (median not reached) compared to their older counterparts (median 25.0 months, P=0.013, hazard ratio=2.08). Treatment intensity, specifically the delivery of a high dose of cytarabine in induction or first consolidation, apparently had a pejorative effect on the outcome of MRD2neg patients compared to standard dose (P=0.048, hazard ratio=1.80), a finding also confirmed by the analysis of data extracted from the literature. The combination of age and treatment intensity allowed us to identify categories of patients, among those who reached a MRD2neg status, characterized by significantly different disease-free survival rate. Our data showed that variables such as age and intensity of treatment administered can influence the predictive value of MRD in patients with acute myeloid leukemia. In addition to underscoring the need for further improvement of MRD analysis, these findings call for a reasoned application of MRD data, as currently available, to modulate consolidation therapy on adequately estimated relapse rates.
Mutations in CCAAT/enhancer binding protein α (
) occur in 5-10% of cases of acute myeloid leukemia.
-double-mutated cases usually bear biallelic N- and C-terminal mutations and are associated with a ...favorable clinical outcome. Identification of
mutants is challenging because of the variety of mutations, intrinsic characteristics of the gene and technical issues. Several screening methods (fragment-length analysis, gene expression array) have been proposed especially for large-scale clinical use; although efficient, they are limited by specific concerns. We investigated the phenotypic profile of blast and maturing bone marrow cell compartments at diagnosis in 251 cases of acute myeloid leukemia. In this cohort, 16 (6.4%) patients had two
mutations, whereas ten (4.0%) had a single mutation. First, we highlighted that the
-double-mutated subset displays recurrent phenotypic abnormalities in all cell compartments. By mutational analysis after cell sorting, we demonstrated that this common phenotypic signature depends on
-double-mutated multi-lineage involvement. From a multidimensional study of phenotypic data, we developed a classifier including ten core and widely available parameters. The selected markers on blasts (CD34, CD117, CD7, CD15, CD65), neutrophil (SSC, CD64), monocytic (CD14, CD64) and erythroid (CD117) compartments were able to cluster
-double-mutated cases. In a validation set of 259 AML cases from three independent centers, our classifier showed excellent performance with 100% specificity and 100% sensitivity. We have, therefore, established a reliable screening method, based upon multidimensional analysis of widely available phenotypic parameters. This method provides early results and is suitable for large-scale detection of
-double-mutated status, allowing gene sequencing to be focused in selected cases.
Summary
A high platelet turnover rate produce a population of immature reticulated platelets (RP) that could confer, despite of antiplatelet drugs, a residual platelet reactivity (RPR) in coronary ...artery disease (CAD) patients. To assess the influence of RP on platelet reactivity in CAD patients on dual antiplatelet therapy we measured RP in 372 patients by using the Sysmex XE-2100 haematology analyzer and platelet function by optical platelet aggregometry (PA) on platelet-rich-plasma induced by 1 mmol arachidonic acid (AA-PA) and 10 μM ADP (ADP-PA). RPR was defined as either AA-PA >20% or ADP-PA >70%. RP were expressed as a percentage of RP of the total optical platelet count (immature platelet fraction; IPF) and as the percentage of RP highly fluorescent (higly fluorescent immature platelet fraction; H-IPF). Moderate but significant positive correlations between PA, IPF, H-IPF, and mean platelet volume (MPV) were found. According to tertiles of IPF, H-IPF and MPV, a significant trend for an increase of platelet aggregation by AA and ADP was evidenced. Furthermore, a significant difference for IPF, H-IPF and MPV between patients with and without RPR was observed. A linear regression analysis showed that IPF, H-IPF and MPV significantly affected PA measured by AA and ADP. At multivariate linear regression analysis these associations were confirmed. Moreover, a logistic regression analysis demonstrated that IPF, H-IPF and MPV significantly influenced the risk of RPR, and in the multivariate model these results remained significant. This study indicates that a high rate of platelet turnover is a new mechanism associated with platelet reactivity in high risk CAD patients on dual antiplatelet therapy.
Highlights • Fingolimod is a sphingosine 1 phosphate receptor (S1P1) agonist approved for oral treatment of patients with relapsing-remitting multiple sclerosis (MS). Its principal mechanisms of ...action is supposed to be to prevent the encephalitogenic attack by sequestering autoreactive lymphocytes in secondary lymphoid organs via downregulation of their S1P1. • Unexpectedly, clinicians have recently found that patients on fingolimod that discontinue the treatment are at risk of extremely severe MS rebound. Although the immunological basis of this rebound is unexplored, elucidating its molecular mechanisms might be of remarkable clinical relevance. • In the present study, we developed the first mouse model of postfingolimod MS rebound. We found that upon fingolimod withdrawal S1P1 overexpression occurs in lymph node-entrapped lymphocytes, and confers them the ability to massively egress form lymphoid organs and bring about a severe CNS infiltration. We also found that S1P1 overexpression that follows fingolimod discontinuation impairs, in a Akt-dependent manner, the ability of regulatory lymphocytes (Tregs) to counteract CNS autoimmunity. • Overall, data disclose the first molecular and cellular mechanisms that may concur to the pathogenesis of postfingolimod rebound in MS patients, and can be targeted for therapeutic interventions. We believe that these novel findings are of importance to those working in the fields of immunological disorders of the CNS and MS therapy.
•Unimodal TRBC1 pattern correlates with T cell clonality.•A TRBC1-including multiparametric panel improves clonality assessment sensitivity.•Monitoring TRBC1 helps in identifying small T-cell clones ...of uncertain significance.•TRBC1 represents an opportunity to overcome time-consuming diagnostic strategies.
The identification of mature T cell neoplasms by flow cytometry is often challenging, due to overlapping features with reactive T cells and limitations of currently available T cell clonality assays. The description of an antibody specific for one of two mutually exclusive T cell receptor (TCR) β-chain constant regions (TRBC1) provides an opportunity to facilitate the detection of clonal TCRαβ+ T cells based on TRBC-restriction. Here we prospectively analyzed 14 healthy controls and 63 patients with the flow cytometry protocol currently used for suspected T cell neoplasm implemented with immunostaining targeting TRBC1. Specimens were firstly classified in 3 groups based on clinical records data, laboratory findings and immunophenotypic features. T cell clonality was assessed by TCR Vβ repertoire analysis and the new rapid TRBC1 assay. Results showed that TRBC1 unimodal expression was unequivocally associated with samples presenting with immunophenotypic aberrancies. Moreover, we demonstrated that the use of TRBC1 is useful in solving uncertain cases and confirmed the high sensitivity of the method in identifying small T cell clones of uncertain significance (T-CUS). Finally, we found a high degree of concordance (97%) comparing the currently available clonality assessment methods with the proposed new method. In conclusion, our results provided real-life evidence of the utility of TRBC1 introduction in the flow cytometric clonality evaluation for the routine diagnostic work-up of T cell neoplasms.
Germ-line mutation rate has been regarded classically as a fundamental biological parameter, as it affects the prevalence of genetic disorders and the rate of evolution. Somatic mutation rate is also ...an important biological parameter, as it may influence the development and/or the course of acquired diseases, particularly of cancer. Estimates of this parameter have been previously obtained in few instances from dermal fibroblasts and lymphoblastoid cells. However, the methodology required has been laborious and did not lend itself to the analysis of large numbers of samples. We have previously shown that the X-linked gene PIG-A, since its product is required for glycosyl-phosphatidylinositol-anchored proteins to become surface bound, is a good sentinel gene for studying somatic mutations. We now show that by this approach we can accurately measure the proportion of PIG-A mutant peripheral blood granulocytes, which we call mutant frequency, ƒ. We found that the results are reproducible, with a variation coefficient (CV) of 45%. Repeat samples from 32 subjects also had a CV of 44%, indicating that ƒ is a relatively stable individual characteristic. From a study of 142 normal subjects we found that log ƒ is a normally distributed variable; ƒ variability spans a 80-fold range, from less than 1×10⁻⁶ to 37.5×10⁻⁶, with a median of 4.9×10⁻⁶. Unlike other techniques commonly employed in population studies, such as comet assay, this method can detect any kind of mutation, including point mutation, as long as it causes functional inactivation of PIG-A gene. Since the test is rapid and requires only a small sample of peripheral blood, this methodology will lend itself to investigating genetic factors that underlie the variation in the somatic mutation rate, as well as environmental factors that may affect it. It will be also possible to test whether ƒ is a determinant of the risk of cancer.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Relapsed/refractory (R/R) acute myeloid leukemia (AML) is a largely unmet medical need, owing to the lack of standardized, effective treatment approaches, resulting in an overall dismal outcome. The ...only curative option for R/R AML patients is allogeneic hematopoietic stem cell transplantation (HSCT) which is only applicable in a fraction of patients due to the scarce efficacy and high toxicity of salvage regimens. Recently, a number of targeted agents with relatively favorable toxicity profiles have been explored in clinical trials for R/R AML patients. The Bcl-2 inhibitor venetoclax, in combination with hypomethylating agents or low dose cytarabine, has produced impressive results for newly diagnosed AML, while its role in R/R disease is not well defined yet. We retrospectively analyzed the clinical outcomes of 47 R/R AML patients treated with venetoclax-based regimens between March 2018 and December 2020 at our institution. Overall, we report a composite complete response rate of 55% with an overall acceptable toxicity profile. Outcomes were particularly favorable for NPM1 mutated patients, unlike for FLT3-ITD positive patients irrespective of NPM1 status. For patients treated with intention to transplant, the procedure could be finally performed in 54%. These findings suggest a role for venetoclax-based regimens in R/R AML patients and support the design of prospective studies.
Recently, a flow cytometric (FC) based test has been developed for detection of circulating fetal cells to replace the less accurate and reproducible Kleihauer-Betke test.
FC test is easier to ...perform, it can distinguish the origin of fetal cells, but it is expensive and available in highly specialized laboratories. We evaluated the introduction of high-performance liquid chromatography (HPLC) approach as initial screening to identify patients who need an additional FC test to better discriminate the nature of haemoglobin-F (HbF) positive cells.
Blood samples from 130 pregnant women suspected to have fetomaternal haemorrhage were analysed with HPLC and FC methods. The cut-off for HbF HPLC concentration was calculated. Statistical analyses for the evaluation of HPLC as a screening method were performed. The positivity cut-off of HbF to be used as decision-making value to continue the investigation was calculated.
An excellent agreement (R2 > 0.90) was observed between the percentage of HbF obtained by HPLC and the percentage of fetal cells detected by FC. Results obtained from each assay were compared to define the HPLC threshold below which it is not necessary to continue the investigations, confirming the maternal nature of the HbF positive cells detected. Our study demonstrated that a cut-off of 1.0 % HbF obtained by HPLC was associated with the lowest rate of false negative results in our patient cohort.
This study provides a new FMH investigation approach that possibly leads to a reduction in times and costs of the analysis.
•HPLC screening test is cheap and could rapidly exclude fetomaternal bleeding.•Unlike the manual Kleihauer-Betke test, HPLC is a standardized automated approach.•Confirmation by flow cytometry could be required only for HPLC positive results.
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