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Multidrug resistance (MDR) mechanisms in cancer cells are greatly influenced by glutathione transferase P1-1 (hGSTP1-1). The use of synthetic or natural compounds as hGSTP1-1 inhibitors is considered an effective approach to overcome MDR. Nine compounds consisting of coumarin-6-sulfonamide linked to chalcone derivatives were synthesized and evaluated for their ability to inhibit hGSTP1-1. Among the synthetic derivatives, compounds 5g, 5f, and 5a displayed the most potent inhibitory effect, with IC50 values of 12.2 ± 0.5 µΜ, 12.7 ± 0.7 and 16.3 ± 0.6, respectively. Kinetic inhibition analysis of the most potent molecule, 5g, showed that it behaves as a mixed-type inhibitor of the target enzyme. An in vitro cytotoxicity assessment of 5a, 5f, and 5g against the human prostate cancer cell lines DU-145 and PC3, as well as the breast cancer cell line MCF-7, demonstrated that compound 5g exhibited the most pronounced cytotoxic effect on all tested cell lines. Molecular docking studies were performed to predict the structural and molecular determinants of 5g, 5f, and 5a binding to hGSTP1-1. In agreement with the experimental data, the results revealed that 5g exhibited the lowest docking score among the three studied inhibitors as a consequence of shape complementarity, governed by van der Waals, hydrogen bonds and a π-π stacking interaction. These findings suggest that coumarin-chalcone hybrids offer new perspectives for the development of safe and efficient natural product-based sensitizers that can target hGSTP1-1 for anticancer purposes.
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Cumarínicos , Glutationa S-Transferase pi , Simulação de Acoplamento Molecular , Sulfonamidas , Humanos , Cumarínicos/química , Cumarínicos/farmacologia , Glutationa S-Transferase pi/antagonistas & inibidores , Glutationa S-Transferase pi/metabolismo , Sulfonamidas/química , Sulfonamidas/farmacologia , Linhagem Celular Tumoral , Chalcona/química , Chalcona/farmacologia , Antineoplásicos/farmacologia , Antineoplásicos/química , Chalconas/química , Chalconas/farmacologia , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/química , Células MCF-7RESUMO
Exposure to sublethal stresses related to food-processing may induce a heterogenous mixture of cells that co-exist, comprising healthy, sublethally injured, dormant and dead cells. Heterogeneity in survival capacity and dormancy of single cells may impede the detection of foodborne pathogens. In this study, we exposed Listeria monocytogenes Scott A strain, to peracetic acid (PAA; 20-40 ppm) and to acidic conditions (hydrochloric (HCl) and acetic (AA) acid, adjusted to pH 2.7-3.0, to evaluate the resuscitation capacity and outgrowth kinetics of metabolically active cells in two different media. Injury and the viable-but-non-culturable (VBNC) status of cells were assessed by flow cytometry using CFDA (metabolically active) and PI (dead) staining. Stressed CFDA+PI- cells were sorted on Tryptic Soy (TS) Agar or in TS broth, both supplemented with 0.6 % Yeast Extract (TSAYE or TSBYE), to evaluate culturability. Resuscitation capacity of CFDA+PI-sorted cells (10 events/well) was monitored by visual inspection on TSAYE and by optical density measurement in TSBYE for 5 days. Sorting of L. monocytogenes viable cells (CFDA+PI-) in Ringer's solution on TSAYE and TSBYE showed 100 % recovery in both media (control condition), while the mean lag time in TSBYE was 9.6 h. Treatment with 20 ppm PAA for 90 and 180 min resulted in 74.79 % and 85.82 % of non-culturable cells in TSBYE and increased the average lag time to 41.7 h and 43.8 h, respectively, compared to the control (9.6 h). The longest average lag time (79.5 h) was detected after treatment with 30 ppm PAA for 90 min, while at the same condition sorting of CFDA+PI- cells resulted in 95.05 % and 93.94 % non-culturable cells on TSAYE and TSBYE, respectively. The highest percentage of wells with non-culturable cells (96.17 %) was detected on TSAYE after treatment with 40 ppm PAA for 30 min. Fractions of VBNC cells were detected in TSBYE after treatment with HCl pH 3.0 for 60 and 240 min, and in TSAYE and TSBYE after exposure to AA pH 2.7. Treatment with AA pH 2.7 for 150-300 min increased the range of recorded lag time values compared to 60 min, from 8.6 h up to 13.3 h, as well as the mean lag times in TSBYE. Modelling of the outgrowth kinetics comparing the two types of stress (oxidative vs acid) and the two systems of growth (colonial vs planktonic) revealed that low starting concentrations hindered the detection of viable L. monocytogenes cells, either due to VBNC induction or cell heterogeneity.
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Microbiologia de Alimentos , Listeria monocytogenes , Listeria monocytogenes/crescimento & desenvolvimento , Viabilidade Microbiana , Ácido Peracético/farmacologia , Ácido Acético/farmacologia , Concentração de Íons de Hidrogênio , Ácido Clorídrico/farmacologia , Contagem de Colônia Microbiana , Meios de Cultura/química , Estresse Fisiológico , Manipulação de Alimentos/métodosRESUMO
There is growing interest in multiple myeloma (MM) circulating tumor cells (CTCs), but their rareness in peripheral blood (PB) and inconsistency in cutoffs question their clinical utility. Herein, we applied next-generation flow cytometry in 550 bone marrow (BM) and matched PB samples to define an optimal CTC cutoff for both transplant-eligible and transplant-ineligible newly diagnosed MM (NDMM) patients. Deep phenotyping was performed to investigate unique microenvironmental features associated with CTC dissemination. CTCs were detected in 90% of patients (median 0.01%; range: 0.0002%-12.6%) and increased levels associated with adverse features. Correlations were observed between high CTC percentages and a diffused MRI pattern, a distinct BM composition characterized by altered B-cell differentiation together with an expansion of effector cells and tumor-associated macrophages, as well as a greater phenotypic dissimilarity between BM and PB clonal cells. Progression-free survival (PFS) and overall survival (OS) gradually worsened with each logarithmic increment of CTCs. Conversely, NDMM patients without CTCs showed unprecedented outcomes, with 5-year PFS and OS rates of 83% and 97%, respectively. A cutoff of 0.02% CTCs was independent of the ISS, LDH, and cytogenetics in a multivariate analysis of risk factors for PFS. The 0.02% CTC cutoff synergized with the MGUS-like phenotype and the R-ISS for improving the risk stratification systems. MRD negativity was less frequent if CTCs were ≥0.02% at diagnosis, but whenever achieved, the poor prognosis of these patients was abrogated. This study shows the clinical utility of CTC assessment in MM and provides evidence toward a consensus cutoff for risk stratification.
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Mieloma Múltiplo , Células Neoplásicas Circulantes , Humanos , Mieloma Múltiplo/patologia , Mieloma Múltiplo/sangue , Mieloma Múltiplo/diagnóstico , Mieloma Múltiplo/mortalidade , Células Neoplásicas Circulantes/patologia , Masculino , Feminino , Pessoa de Meia-Idade , Idoso , Adulto , Idoso de 80 Anos ou mais , Prognóstico , Medula Óssea/patologiaRESUMO
Preclinical and clinical data demonstrate synergy between belantamab mafodotin (belamaf) and immunomodulatory drugs with limited overlapping toxicities. We investigated the safety and efficacy of belamaf with lenalidomide 25 mg on days 1-21 every 28 days and dexamethasone 40 mg weekly (belamaf-Rd) in transplant-ineligible patients with newly diagnosed multiple myeloma. Thirty-six patients (median age, 72.5 years) were randomized to receive belamaf at three different doses (2.5, 1.9, or 1.4 mg/kg) every 8 weeks. The dosing schedule was extended to every 12 weeks to mitigate ocular toxicity. Most common grade ≥3 adverse events were fatigue (n=21, 58.3%), rash (n=6, 16.7%), diarrhea (n=8, 22.2%) and COVID-19 (n=5, 13.9%). Grade 3-4 ocular adverse events, comprising visual acuity decline from baseline and/or keratopathy, were reported in 39/216 (18.1%), 33/244 (13.5%), and 26/207 (12.6%) ophthalmological assessments in the 2.5, 1.9, and 1.4 mg/kg cohorts, respectively. Importantly, grade 3-4 keratopathy was identified in 9/216 (4.2%), 1/244 (0.4%) and 1/207(0.5%) assessments. Most patients (32/36, 88.9%) were treated with the extended, every-12-week schedule, during which 40, 33 and 16 doses were withheld due to ocular adverse events in the 2.5, 1.9, and 1.4 mg/kg cohorts, respectively. Overall, the rates of very good partial response and better and complete response and better were 83.3% and 52.8%, respectively, without significant differences among cohorts. Over a median follow-up of 20.3 months no disease progression was reported; six patients discontinued treatment due to infection-related death (4 cases of COVID-19, 2 cases of pneumonia) and one patient withdrew consent. Based on the toxicity/efficacy balance, the recommended phase II dose was 1.9 mg/kg every 8 weeks, extended to every 12 weeks because of toxicity. In conclusion, Belamaf-Rd, with the extended schedule for belamaf, showed important clinical activity and a significant improvement of ocular adverse events with minimal impact on vision-related functioning in an elderly, non-transplant eligible population.
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Protocolos de Quimioterapia Combinada Antineoplásica , Dexametasona , Lenalidomida , Mieloma Múltiplo , Humanos , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/diagnóstico , Lenalidomida/administração & dosagem , Lenalidomida/efeitos adversos , Lenalidomida/uso terapêutico , Idoso , Masculino , Feminino , Dexametasona/administração & dosagem , Dexametasona/efeitos adversos , Dexametasona/uso terapêutico , Pessoa de Meia-Idade , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Idoso de 80 Anos ou mais , SARS-CoV-2 , Resultado do Tratamento , COVID-19 , Anticorpos Monoclonais HumanizadosRESUMO
Inflammasomes are multimeric protein complexes, sensors of intracellular danger signals, and crucial components of the innate immune system, with the NLRP3 inflammasome being the best characterized among them. The increasing scientific interest in the mechanisms interconnecting inflammation and tumorigenesis has led to the study of the NLRP3 inflammasome in the setting of various neoplasms. Despite a plethora of data regarding solid tumors, NLRP3 inflammasome's implication in the pathogenesis of hematological malignancies only recently gained attention. In this review, we investigate its role in normal lymphopoiesis and lymphomagenesis. Considering that lymphomas comprise a heterogeneous group of hematologic neoplasms, both tumor-promoting and tumor-suppressing properties were attributed to the NLRP3 inflammasome, affecting neoplastic cells and immune cells in the tumor microenvironment. NLRP3 inflammasome-related proteins were associated with disease characteristics, response to treatment, and prognosis. Few studies assess the efficacy of NLRP3 inflammasome therapeutic targeting with encouraging results, though most are still at the preclinical level. Further understanding of the mechanisms regulating NLRP3 inflammasome activation during lymphoma development and progression can contribute to the investigation of novel treatment approaches to cover unmet needs in lymphoma therapeutics.
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Inflamassomos , Linfoma , Humanos , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Inflamação/metabolismo , Linfoma/etiologia , Linfoma/terapia , Microambiente TumoralRESUMO
Obesity in adolescence is associated with significant morbidity and predisposes adolescents to the development of cardiovascular disease (CVD). Although a number of traditional CVD risk factors have been identified in youth, limited data exist regarding non-traditional CVD risk factors. In 89 adolescents with metabolic syndrome (MetS), with 60 age-, gender-, and BMI-matched controls, we determined the non-traditional CVD risk factors (hs-CRP, TG/HDL ratio, ApoB/ApoA1 ratio, NAFLD) in order to investigate whether they may be used as biomarkers for predicting future CVD, and we evaluated their response to the implementation of a multidisciplinary, personalized, lifestyle intervention program for 1 year. We demonstrated that the TG/HDL ratio, IL-2, IL-6, IL-17A, and INF-γ were significantly increased in subjects with MetS than in controls, and may be used as biomarkers to predict future CVD. Subjects with MetS had an increased mean carotid intima-media thickness (cIMT) and prevalence of NAFLD than the controls, while the prevalence of NAFLD correlated strongly with cIMT and IL-6 concentrations. Most of the non-traditional cardiovascular risk factors improved following the implementation of a lifestyle intervention program. These findings indicate that adolescents with MetS may have a greater risk for developing atherosclerosis early in life, while early lifestyle intervention is crucial for preventing the arteriosclerotic process in youth.
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Doenças Cardiovasculares , Síndrome Metabólica , Hepatopatia Gordurosa não Alcoólica , Obesidade Infantil , Adolescente , Humanos , Síndrome Metabólica/complicações , Síndrome Metabólica/epidemiologia , Obesidade Infantil/complicações , Obesidade Infantil/epidemiologia , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/complicações , Hepatopatia Gordurosa não Alcoólica/epidemiologia , Hepatopatia Gordurosa não Alcoólica/complicações , Fatores de Risco , Espessura Intima-Media Carotídea , Interleucina-6 , Fatores de Risco de Doenças Cardíacas , BiomarcadoresRESUMO
Erythropoietin-producing hepatocellular carcinoma receptors (EPHs) represent the largest family of receptor tyrosine kinases (RTKs). EPH interaction with ephrins, their membrane-bound ligands, holds a pivotal role in embryonic development, while, though less active, it is also implicated in various physiological functions during adult life. In normal hematopoiesis, different patterns of EPH/ephrin expression have been correlated with hematopoietic stem cell (HSC) maintenance and lineage-committed hematopoietic progenitor cell (HPC) differentiation, as well as with the functional properties of their mature offspring. Research in the field of hematologic malignancies has unveiled a rather complex involvement of the EPH/ephrinsignaling pathway in the pathophysiology of these neoplasms. Aberrations in genetic, epigenetic, and protein levels have been identified as possible players implicated both in tumor progression and suppression, while correlations have also been highlighted regarding prognosis and response to treatment. Initial efforts to therapeutically target the EPH/ephrin axis have been undertaken in the setting of hematologic neoplasia but are mainly confined to the preclinical level. To this end, deciphering the complexity of this signaling pathway both in normal and malignant hematopoiesis is necessary.
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Bladder cancer (BLCA) is the sixth most common type of cancer and has a dismal prognosis if diagnosed late. To identify treatment options for BLCA, we systematically evaluated data from the Broad Institute DepMap project. We found that urothelial BLCA cell lines are among the most sensitive to microtubule assembly inhibition by paclitaxel treatment. Strikingly, we revealed that the top dependencies in BLCA cell lines include genes encoding proteins involved in microtubule assembly. This highlights the importance of microtubule network dynamics as a major vulnerability in human BLCA. In cancers such as ovarian and breast, where paclitaxel is the gold standard of care, resistance to paclitaxel treatment has been linked to p53-inactivating mutations. To study the response of BLCA to microtubule assembly inhibition and its mechanistic link with the mutational status of the p53 protein, we treated a collection of BLCA cell lines with a dose range of paclitaxel and performed a detailed characterization of the response. We discovered that BLCA cell lines are significantly sensitive to low concentrations of paclitaxel, independently of their p53 status. Paclitaxel induced a G2/M cell cycle arrest and growth inhibition, followed by robust activation of apoptosis. Most importantly, we revealed that paclitaxel triggered a robust DNA-damage response and apoptosis program without activating the p53 pathway. Integration of transcriptomics, epigenetic, and dependency data demonstrated that the response of BLCA to paclitaxel is independent of p53 mutational signatures but strongly depends on the expression of DNA repair genes. Our work highlights urothelial BLCA as an exceptional candidate for paclitaxel treatment. It paves the way for the rational use of a combination of paclitaxel and DNA repair inhibitors as an effective, novel therapeutic strategy.
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BACKGROUND: Chemotherapy (CT) is central to the treatment of triple negative breast cancer (TNBC), but drug toxicity and resistance place strong restrictions on treatment regimes. Fasting sensitizes cancer cells to a range of chemotherapeutic agents and also ameliorates CT-associated adverse effects. However, the molecular mechanism(s) by which fasting, or short-term starvation (STS), improves the efficacy of CT is poorly characterized. METHODS: The differential responses of breast cancer or near normal cell lines to combined STS and CT were assessed by cellular viability and integrity assays (Hoechst and PI staining, MTT or H2DCFDA staining, immunofluorescence), metabolic profiling (Seahorse analysis, metabolomics), gene expression (quantitative real-time PCR) and iRNA-mediated silencing. The clinical significance of the in vitro data was evaluated by bioinformatical integration of transcriptomic data from patient data bases: The Cancer Genome Atlas (TCGA), European Genome-phenome Archive (EGA), Gene Expression Omnibus (GEO) and a TNBC cohort. We further examined the translatability of our findings in vivo by establishing a murine syngeneic orthotopic mammary tumor-bearing model. RESULTS: We provide mechanistic insights into how preconditioning with STS enhances the susceptibility of breast cancer cells to CT. We showed that combined STS and CT enhanced cell death and increased reactive oxygen species (ROS) levels, in association with higher levels of DNA damage and decreased mRNA levels for the NRF2 targets genes NQO1 and TXNRD1 in TNBC cells compared to near normal cells. ROS enhancement was associated with compromised mitochondrial respiration and changes in the metabolic profile, which have a significant clinical prognostic and predictive value. Furthermore, we validate the safety and efficacy of combined periodic hypocaloric diet and CT in a TNBC mouse model. CONCLUSIONS: Our in vitro, in vivo and clinical findings provide a robust rationale for clinical trials on the therapeutic benefit of short-term caloric restriction as an adjuvant to CT in triple breast cancer treatment.
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Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Neoplasias de Mama Triplo Negativas , Animais , Camundongos , Humanos , Dieta Redutora , Espécies Reativas de Oxigênio , ObesidadeRESUMO
BACKGROUND: Based on the fact that coronavirus disease 2019 (COVID-19) is still spreading despite worldwide vaccine administration, there is an imperative need to understand the underlying mechanisms of vaccine-induced interindividual immune response variations. METHODS: We compared humoral and cellular immune responses in 127 individuals vaccinated with either BNT162b2, mRNA-1273, or ChAdOx1-nCoV-19 vaccine. RESULTS: Both mRNA vaccines induced faster and stronger humoral responses as assessed by high spike- and RBD-specific antibody titers and neutralizing efficacy in comparison to ChAdOx1-nCoV-19 vaccine. At 7 months postvaccination, a decreasing trend in humoral responses was observed, irrespective of the vaccine administered. Correlation analysis between anti-S1 IgG and interferon- (IFN-) production unveiled a heterogeneous immune profile among BNT162b2-vaccinated individuals. Specifically, vaccination in the high-responder group induced sizable populations of polyfunctional memory CD4 helper T cells (TH1), follicular helper T cells (TFH), and T cells with features of stemness (TSCM), along with high neutralizing antibody production that persisted up to 7 months. In contrast, low responders were characterized by significantly lower antibody titers and memory T cells and a considerably lower capacity for interleukin-2 and IFN- production. CONCLUSIONS: We identified that long-term humoral responses correlate with the individuals ability to produce antigen-specific persistent memory T-cell populations.
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COVID-19 , SARS-CoV-2 , Humanos , Vacina BNT162 , COVID-19/prevenção & controle , Subpopulações de Linfócitos T , Anticorpos Neutralizantes , Anticorpos Antivirais , VacinaçãoRESUMO
BACKGROUND/OBJECTIVE: Older age and male sex have been consistently found to be associated with dismal outcomes among COVID-19 infected patients. In contrast, premenopausal females present the lowest mortality among adults infected by SARS-CoV-2. The goal of the present study was to investigate whether peripheral blood type I interferon (IFN) signature and interleukin (IL)-6 serum levels -previously shown to contribute to COVID-19-related outcomes in hospitalized patients- is shaped by demographic contributors among COVID-19 convalescent individuals. PATIENTS AND METHODS: Type I IFN-inducible genes in peripheral blood, as well as serum IL-6 levels were quantified in 61 COVID-19 convalescent healthy individuals (34 females, 27 males; age range 18-70 years, mean 35.7 ± 15.9 years) who recovered from COVID-19 without requiring hospitalization within a median of 3 months prior to inclusion in the present study. Among those, 17 were older than 50 years (11 males, 6 females) and 44 equal to or less than 50 years (16 males, 28 females). Expression analysis of type I IFN-inducible genes (MX-1, IFIT-1, IFI44) was performed by real time PCR and a type I IFN score, reflecting type I IFN peripheral activity, was calculated. IL-6 and C-reactive protein levels were determined by a commercially available ELISA. RESULTS: COVID-19 convalescent individuals older than 50 years exhibited significantly decreased peripheral blood type I IFN scores along with significantly increased IL-6 serum levels compared to their younger counterparts less than 50 years old (5.4 ± 4.3 vs 16.8 ± 24.7, p = 0.02 and 10.6 ± 16.9 vs 2.9 ± 8.0 ng/L, p = 0.03, respectively). Following sex stratification, peripheral blood type I IFN score was found to be significantly higher in younger females compared to both younger and older males (22.9 ± 29.2 vs 6.3 ± 4.6 vs 4.5 ± 3.7, p = 0.01 and p = 0.002, respectively). Regarding IL-6, an opposite pattern was observed, with the highest levels being detected among older males and the lowest levels among younger females (11.6 ± 18.9 vs 2.5 ± 7.8 ng/L, p = 0.03). CONCLUSION: Constitutive higher type I IFN responses and dampened IL-6 production observed in younger women of premenopausal age, along with lower type I IFN responses and increased IL-6 levels in older males, could account for the discrete clinical outcomes seen in the two population groups, as consistently revealed in COVID-19 epidemiological studies.
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COVID-19 , Interferon Tipo I , Adulto , Idoso , Pré-Escolar , Feminino , Hospitalização , Humanos , Lactente , Interleucina-6 , Masculino , Pessoa de Meia-Idade , SARS-CoV-2RESUMO
The new and increasingly studied concept of immunogenic cell death (ICD) revealed a previously unknown perspective of the various regulated cell death (RCD) modalities, elucidating their immunogenic properties and rendering obsolete the notion that immune stimulation is solely the outcome of necrosis. A distinct characteristic of ICD is the release of danger-associated molecular patterns (DAMPs) by dying and/or dead cells. Thus, several members of the DAMP family, such as the well-characterized heat shock proteins (HSPs) HSP70 and HSP90, the high-mobility group box 1 protein and calreticulin, and the thymic polypeptide prothymosin α (proTα) and its immunoreactive fragment proTα(100-109), are being studied as potential diagnostic tools and/or possible therapeutic agents. Here, we present the basic aspects and mechanisms of both ICD and other immunogenic RCD forms; denote the role of DAMPs in ICD; and further exploit the relevance of human proTα and proTα(100-109) in ICD, highlighting their possible clinical applications. Furthermore, we present the preliminary results of our in vitro studies, which show a direct correlation between the concentration of proTα/proTα(100-109) and the levels of cancer cell apoptosis, induced by anticancer agents and γ-radiation.
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Morte Celular Imunogênica , Timosina , Alarminas/metabolismo , Biomarcadores , Humanos , Imunidade , Peptídeos , Precursores de Proteínas , Timosina/análogos & derivados , Timosina/farmacologiaRESUMO
Plasma cell leukemia (PCL) is a rare and aggressive plasma cell dyscrasia that may appear as de-novo leukemia (pPCL) or on the basis of a pre-existing multiple myeloma (MM), called secondary plasma cell leukemia (sPCL). In this prospective study, we have applied a broad panel of FISH probes in 965 newly diagnosed MM (NDMM) and 44 PCL cases of both types to reveal the particular cytogenetic differences among the three plasma cell dyscrasias. In order to evaluate the frequency and patterns of clonal evolution, the same FISH panel was applied both at diagnosis and at the time of first relapse for 81 relapsed MM patients and both at MM diagnosis and during sPCL transformation for the 19 sPCL cases described here. pPCL was characterized by frequent MYC translocations and t(11;14) with a 11q13 breakpoint centered on the MYEOV gene, not commonly seen in MM. sPCL had a higher number of FISH abnormalities and was strongly associated with the presence of del(17p13), either acquired at the initial MM stage or as a newly acquired lesion upon leukemogenesis in the context of the apparent clonal evolution observed in sPCL. In clinical terms, sPCL showed a shorter overall survival than pPCL with either standard or high-risk (t(4;14) and/or t(14;16) and/or del(17p13) and/or ≥3 concomitant aberrations) abnormalities (median 5 months vs. 21 and 11 months respectively, p < 0.001), suggesting a prognostic stratification based on cytogenetic background. These observations proved relevant in the NDMM setting, where higher levels of circulating plasma cells (CPCs) were strongly associated with high-risk cytogenetics (median frequency of CPCs: 0.11% of peripheral blood nucleated cells for high-risk vs. 0.007% for standard-risk NDMM, p < 0.0001). Most importantly, the combined evaluation of CPCs (higher or lower than a cut-off of 0.03%), together with patients' cytogenetic status, could be used for an improved prognostic stratification of NDMM patients.
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BACKGROUND: Members of the α-thymosin family have long been studied for their immunostimulating properties. Among them, the danger-associated molecular patterns (DAMPs) prothymosin α (proTα) and its C-terminal decapeptide proTα(100-109) have been shown to act as immunomodulators in vitro, due to their ability to promote T helper type 1 (Th1) responses. Recently, we verified these findings in vivo, showing that both proTα and proTα(100-109) enhance antitumor-reactive T cell-mediated responses. METHODS: In view of the eventual use of proTα and proTα(100-109) in humans, we investigated their safety profile in silico, in human leukocytes and cancer cell lines in vitro, and in immunocompetent mice in vivo, in comparison to the proTα derivative thymosin alpha 1 (Τα1), a 28-mer peptide extensively studied for its safety in clinical trials. RESULTS: In silico prediction via computational tools showed that all three peptide sequences likely are non-toxic or do not induce allergic regions. In vitro, pro- Tα, proTα(100-109) and Tα1 did not affect the viability of human cancer cell lines and healthy donor-derived leukocytes, did not promote apoptosis or alter cell cycle distribution. Furthermore, mice injected with proTα, proTα(100-109) and Tα1 at doses equivalent to the suggested dose regimen of Tα1 in humans, did not show signs of acute toxicity, whereas proTα and proTα(100-109) increased the levels of proinflammatory and Th1- type cytokines in their peripheral blood. CONCLUSION: Our preliminary findings suggest that proTα and proTα(100-109), even at high concentrations, are non-toxic in vitro and in an acute toxicity model in vivo; moreover, we show that the two peptides retain their immunomodulatory properties in vivo and, eventually, could be considered for therapeutic use in humans.
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Neoplasias , Timosina , Humanos , Camundongos , Animais , Timosina/toxicidade , Peptídeos/uso terapêutico , Citocinas , Fatores Imunológicos/uso terapêutico , Neoplasias/tratamento farmacológicoRESUMO
Blood donor genetics and lifestyle affect the quality of red blood cell (RBC) storage. Heterozygotes for beta thalassemia (bThal+) constitute a non-negligible proportion of blood donors in the Mediterranean and other geographical areas. The unique hematological profile of bThal+ could affect the capacity of enduring storage stress, however, the storability of bThal+ RBC is largely unknown. In this study, RBC from 18 bThal+ donors were stored in the cold and profiled for primary (hemolysis) and secondary (phosphatidylserine exposure, potassium leakage, oxidative stress) quality measures, and metabolomics, versus sex- and age-matched controls. The bThal+ units exhibited better levels of storage hemolysis and susceptibility to lysis following osmotic, oxidative and mechanical insults. Moreover, bThal+ RBC had a lower percentage of surface removal signaling, reactive oxygen species and oxidative defects to membrane components at late stages of storage. Lower potassium accumulation and higher uratedependent antioxidant capacity were noted in the bThal+ supernatant. Full metabolomics analyses revealed alterations in purine and arginine pathways at baseline, along with activation of the pentose phosphate pathway and glycolysis upstream to pyruvate kinase in bThal+ RBC. Upon storage, substantial changes were observed in arginine, purine and vitamin B6 metabolism, as well as in the hexosamine pathway. A high degree of glutamate generation in bThal+ RBC was accompanied by low levels of purine oxidation products (IMP, hypoxanthine, allantoin). The bThal mutations impact the metabolism and the susceptibility to hemolysis of stored RBC, suggesting good post-transfusion recovery. However, hemoglobin increment and other clinical outcomes of bThal+ RBC transfusion deserve elucidation by future studies.
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Talassemia beta , Preservação de Sangue , Transfusão de Eritrócitos , Eritrócitos/metabolismo , Hemólise , Humanos , Talassemia beta/genética , Talassemia beta/metabolismoRESUMO
The 4th Summer School in Immuno-Oncology was held from July 1st-July 3rd as a web meeting. Many eminent researchers and leading oncologists from Europe and the USA working on basic, translational and clinical cancer research participated, presented, and discussed the most recent advances in cancer immunology and immunotherapy. Besides sharing the newest information in the field of cancer immunology and immunotherapy, the meeting also focused on the actual translation of new knowledge acquired in the lab to the clinical setting; particular emphasis was given to the mode of action of novel therapeutic modalities and to biomarkers helpful for treatment decision-making, as well as to means that may improve cancer immunotherapeutic protocols used for the treatment of a variety of malignancies. The main topics presented by the speakers included: (1) mechanisms of tumor immune evasion and resistance; (2) host-tumor interactions and means to regulate antitumor immunity; (3) exploitation of new biomarkers and tumor or immune signatures able to potentially guide therapeutic interventions; (4) emerging therapeutic modalities for cancer treatment and specific immunotherapeutics for thoracic, genito-urinary, gastrointestinal, skin and breast cancers; and (5) innovative treatment options and alternatives to minimize the toxic adverse events of cancer immunotherapy.
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Neoplasias , Grécia , Humanos , Imunoterapia , Oncologia , Neoplasias/terapia , Instituições AcadêmicasRESUMO
High-dose chemotherapy with autologous stem cell support (ASCT) is the standard of care for eligible newly diagnosed Multiple Myeloma (MM) patients. Stem cell graft contamination by aberrant plasma cells (APCs) has been considered a possible predictive marker of subsequent clinical outcome, but the limited reports to date present unclear conclusions. We prospectively estimated the frequency of graft contamination using highly sensitive next-generation flow cytometry and evaluated its clinical impact in 199 myeloma patients who underwent an ASCT. Contamination (con+) was detected in 79/199 patients at a median level 2 × 10-5. Its presence and levels were correlated with response to induction treatment, with 94%, 71% and 43% achieving CR, VGPR and PR, respectively. Importantly, con+ grafts conferred 2-fold and 2.8-fold higher patient-risk of not achieving or delaying reaching CR (4 vs. 11 months) and MRD negativity (5 vs. 18 months) post ASCT, respectively. Our data also provide evidence of a potentially skewed bone marrow (BM) reconstitution due to unpurged grafts, since con+ derived BM had significantly higher prevalence of memory B cells. These data, together with the absence of significant associations with baseline clinical features, highlight graft contamination as a potential biomarker with independent prognostic value for deeper responses, including MRD negativity. Longer follow-up will reveal if this corresponds to PFS or OS advantage.
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Between June and November 2020, we assessed plasma antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein in 4996 participants (aged 18-82 years, 34.5% men) from the National and Kapodistrian University of Athens. The weighted overall prevalence was 1.6% and monthly prevalence correlated with viral RNA-confirmed SARS-CoV-2 infections in Greece, in the same period. Notably, 49% of seropositive cases reported no history of SARS-CoV-2 infection-related clinical symptoms and 33% were unsuspected of their previous infection. Additionally, levels of anti-SARS-CoV-2 antibodies against the spike-protein receptor-binding domain were similar between symptomatic and asymptomatic individuals, irrespective of age and gender. Using Food and Drug Administration Emergency Use Authorization-approved assays, these results support the need for such studies on pandemic evaluation and highlight the development of robust humoral immune responses even among asymptomatic individuals. The high percentage of unsuspected/asymptomatic active cases, which may contribute to community transmission for more days than that of cases who are aware and self-isolate, underscores the necessity of measures across the population for the efficient control of the pandemic.
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Melanoma is classified among the most notoriously aggressive human cancers. Despite the recent progress, due to its propensity for metastasis and resistance to therapy, novel biomarkers and oncogenic molecular drivers need to be promptly identified for metastatic melanoma. Hence, by employing nano liquid chromatography-tandem mass spectrometry deep proteomics technology, advanced bioinformatics algorithms, immunofluorescence, western blotting, wound healing protocols, molecular modeling programs, and MTT assays, we comparatively examined the respective proteomic contents of WM115 primary (n = 3955 proteins) and WM266-4 metastatic (n = 6681 proteins) melanoma cells. It proved that WM115 and WM266-4 cells have engaged hybrid epithelial-to-mesenchymal transition/mesenchymal-to-epithelial transition states, with TGF-ß controlling their motility in vitro. They are characterized by different signatures of SOX-dependent neural crest-like stemness and distinct architectures of the cytoskeleton network. Multiple signaling pathways have already been activated from the primary melanoma stage, whereas HIF1α, the major hypoxia-inducible factor, can be exclusively observed in metastatic melanoma cells. Invasion-metastasis cascade-specific sub-routines of activated Caspase-3-triggered apoptosis and LC3B-II-dependent constitutive autophagy were also unveiled. Importantly, WM115 and WM266-4 cells exhibited diverse drug response profiles, with epirubicin holding considerable promise as a beneficial drug for metastatic melanoma clinical management. It is the proteome navigation that enables systemic biomarkering and targeted drugging to open new therapeutic windows for advanced disease.
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Persisting alterations and unique immune signatures have been previously detected in the peripheral blood of convalescent plasma (CP) donors at approximately two months after initial SARS-CoV-2 infection. This article presents the results on the sequential analysis of 47 CP donors at a median time of eight months (range 7.5-8.5 months) post infection, as assessed by flow cytometry. Interestingly, our results show a significant variation of the relevant immune subset composition among CP donors. Regarding innate immunity, both non-classical monocytes, and CD11b- granulocytes had fully recovered at eight months post COVID-19 infection. Intermediate monocytes and natural killer (NK) cells had already been restored at the two-month evaluation and remained stable. Regarding adaptive immunity, the COVID-19-related skewed Th1 and Th2 cell polarization remained at the same levels as in two months. However, low levels of total B cells were detected even after eight months from infection. A persisting reduction of CD8+ Tregs and changes in the NKT cell compartment were also remarkable. CP donors present with a unique immune landscape at eight months post COVID-19 infection, which is characterized by the notable restoration of the components of innate immunity along with a persisting imprint of SARS-CoV-2 in cells of the adaptive immunity.