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BACKGROUND: Cutaneous leishmaniasis (CL) caused by Leishmania (Viannia) braziliensis is associated with an inflammatory response. Granzyme (GzmB) and IL-1ß play a key role in the pathology. Meglumine antimoniate (MA) is the first-choice drug for the treatment of CL, but therapy failure is observed in up to 50% of the cases. The protein, rSm29 of Schistosoma mansoni, down modulates pro-inflammatory cytokine production. We evaluate if the combination of topical rSm29 plus MA increases the cure rate of CL. METHODS: In this randomized clinical trial, 91 CL patients were allocated in 3 groups. All cases received MA (20mg/Kg/weight) for 20 days. Group 1 used topical rSm29 (10ug), group 2 a placebo topically applied, and group 3 received only MA. RESULTS: The cure rate on day 90 was 71% in subjects treated with rSm29 plus MA, and 43% in patients who received MA plus placebo or MA alone (P<0.05). There was a decrease in GzmB and an increase in IFN-γ (P<0.05) in supernatants of skin biopsies of the lesions obtained on D7 of therapy (P<0.05) in patients who received rSm29. CONCLUSION: rSm29 associated with MA reduces GzmB levels, is more effective than MA alone and decreases CL healing time. CLINICAL TRIALS REGISTRATION: ClinicalTrial.gov under NCT06000514.
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Nanotechnology plays a crucial role in vaccine development and provides the opportunity to design functional nanoparticles (Np) of different compositions, sizes, charges and surface properties for biomedical applications. The present study aims to evaluate a complex coacervate-like Np composed of poly(allylamine hydrochloride) (PAH) and tripolyphosphate (Tpp) as a safe vehicle and adjuvant for systemic vaccines. We investigated the activation of different antigen-presenting cells (APCs) with Np-PAH and its adjuvanticity in Balbc/c and different KO mice that were intraperitoneally immunized with Np-OVA. We found that Np-PAH increased the expression of CD86 and MHCII and promoted the production and secretion of interleukin-1ß (IL-1ß) and IL-18 through the inflammasome NLRP3 when macrophages and dendritic cells were co-incubated with LPS and Np-PAH. We evidenced an unconventional IL-1ß release through the autophagosome pathway. The inhibition of autophagy with 3-methyladenine reduced the LPS/Np-PAH-induced IL-1ß secretion. Additionally, our findings showed that the systemic administration of mice with Np-OVA triggered a significant induction of serum OVA-specific IgG and IgG2a, an increased secretion of IFN-γ by spleen cells, and high frequencies of LT CD4 + IFN-γ + and LT CD8 + IFN-γ + . In conclusion, our findings show that PAH-based Np promoted the inflammasome activation of innate cells with Th1-dependent adjuvant properties, making them valuable for formulating of novel preventive or therapeutic vaccines for infectious and non-infectious diseases.
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Bacillus Calmette-Guérin (BCG) is the first line treatment for bladder cancer and it is also proposed for melanoma immunotherapy. BCG modulates the tumor microenvironment (TME) inducing an antitumor effective response, but the immune mechanisms involved still poorly understood. The immune profile of B16-F10 murine melanoma cells was assessed by infecting these cells with BCG or stimulating them with agonists for different innate immune pathways such as TLRs, inflammasome, cGAS-STING and type I IFN. B16-F10 did not respond to any of those stimuli, except for type I IFN agonists, contrasting with bone marrow-derived macrophages (BMDMs) that showed high production of proinflammatory cytokines. Additionally, we confirmed that BCG is able to infect B16-F10, which in turn can activate macrophages and spleen cells from mice in co-culture experiments. Furthermore, we established a subcutaneous B16-F10 melanoma model for intratumoral BCG treatment and compared wild type mice to TLR2-/-, TLR3-/-, TLR4-/-, TLR7-/-, TLR3/7/9-/-, caspase 1-/-, caspase 11-/-, IL-1R-/-, cGAS-/-, STING-/-, IFNAR-/-, MyD88-/-deficient animals. These results in vivo demonstrate that MyD88 signaling is important for BCG immunotherapy to control melanoma in mice. Also, BCG fails to induce cytokine production in the co-culture experiments using B16-F10 and BMDMs or spleen cells derived from MyD88-/- compared to wild-type (WT) animals. Immunotherapy with BCG was not able to induce the recruitment of inflammatory cells in the TME from MyD88-/- mice, impairing tumor control and IFN-γ production by T cells. In conclusion, MyD88 impacts on both innate and adaptive responses to BCG leading to an efficient antitumor response against melanoma.
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Vacina BCG , Imunoterapia , Melanoma Experimental , Fator 88 de Diferenciação Mieloide , Transdução de Sinais , Animais , Camundongos , Vacina BCG/imunologia , Vacina BCG/uso terapêutico , Linhagem Celular Tumoral , Citocinas/metabolismo , Imunoterapia/métodos , Macrófagos/imunologia , Macrófagos/metabolismo , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium bovis/imunologia , Fator 88 de Diferenciação Mieloide/metabolismo , Fator 88 de Diferenciação Mieloide/genética , Microambiente Tumoral/imunologiaRESUMO
Neutrophils can be beneficial or deleterious during tuberculosis (TB). Based on the expression of MHC-II and programmed death ligand 1 (PD-L1), we distinguished two functionally and transcriptionally distinct neutrophil subsets in the lungs of mice infected with mycobacteria. Inflammatory [MHC-II-, PD-L1lo] neutrophils produced inflammasome-dependent IL-1ß in the lungs in response to virulent mycobacteria and "accelerated" deleterious inflammation, which was highly exacerbated in IFN-γR-/- mice. Regulatory [MHC-II+, PD-L1hi] neutrophils "brake" inflammation by suppressing T-cell proliferation and IFN-γ production. Such beneficial regulation, which depends on PD-L1, is controlled by IFN-γR signaling in neutrophils. The hypervirulent HN878 strain from the Beijing genotype curbed PD-L1 expression by regulatory neutrophils, abolishing the braking function and driving deleterious hyperinflammation in the lungs. These findings add a layer of complexity to the roles played by neutrophils in TB and may explain the reactivation of this disease observed in cancer patients treated with anti-PD-L1.
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Antígeno B7-H1 , Inflamação , Interleucina-1beta , Pulmão , Neutrófilos , Tuberculose , Animais , Antígeno B7-H1/metabolismo , Antígeno B7-H1/genética , Neutrófilos/imunologia , Neutrófilos/metabolismo , Camundongos , Interleucina-1beta/metabolismo , Inflamação/imunologia , Inflamação/metabolismo , Tuberculose/imunologia , Tuberculose/microbiologia , Tuberculose/metabolismo , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/metabolismo , Pulmão/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium tuberculosis/imunologia , Modelos Animais de Doenças , Feminino , HumanosRESUMO
Introduction: Guanylate-binding proteins (GBPs) are produced in response to pro-inflammatory signals, mainly interferons. The most studied cluster of GBPs in mice is on chromosome 3. It comprises the genes for GBP1-to-3, GBP5 and GBP7. In humans, all GBPs are present in a single cluster on chromosome 1. Brucella abortus is a Gram-negative bacterium known to cause brucellosis, a debilitating disease that affects both humans and animals. Our group demonstrated previously that GBPs present on murine chromosome 3 (GBPchr3) is important to disrupt Brucella-containing vacuole and GBP5 itself is important to Brucella intracellular LPS recognition. In this work, we investigated further the role of GBPs during B. abortus infection. Methods and results: We observed that all GBPs from murine chromosome 3 are significantly upregulated in response to B. abortus infection in mouse bone marrow-derived macrophages. Of note, GBP5 presents the highest expression level in all time points evaluated. However, only GBPchr3-/- cells presented increased bacterial burden compared to wild-type macrophages. Brucella DNA is an important Pathogen-Associated Molecular Pattern that could be available for inflammasome activation after BCV disruption mediated by GBPs. In this regard, we observed reduced IL-1ß production in the absence of GBP2 or GBP5, as well as in GBPchr3-/- murine macrophages. Similar result was showed by THP-1 macrophages with downregulation of GBP2 and GBP5 mediated by siRNA. Furthermore, significant reduction on caspase-1 p20 levels, LDH release and Gasdermin-D conversion into its mature form (p30 N-terminal subunit) was observed only in GBPchr3-/- macrophages. In an in vivo perspective, we found that GBPchr3-/- mice had increased B. abortus burden and higher number of granulomas per area of liver tissue, indicating increased disease severity. Discussion/conclusion: Altogether, these results demonstrate that although GBP5 presents a high expression pattern and is involved in inflammasome activation by bacterial DNA in macrophages, the cooperation of multiple GBPs from murine chromosome 3 is necessary for full control of Brucella abortus infection.
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Brucelose , Proteínas de Ligação ao GTP , Animais , Camundongos , Brucella abortus/genética , Brucelose/microbiologia , Proteínas de Transporte/metabolismo , DNA Bacteriano , Inflamassomos/genética , Inflamassomos/metabolismo , Proteínas de Ligação ao GTP/genéticaRESUMO
The bacillus Calmette-Guérin (BCG) is an attenuated bacterium derived from virulent Mycobacterium bovis. It is the only licensed vaccine used for preventing severe forms of tuberculosis in children. Besides its specific effects against tuberculosis, BCG administration is also associated with beneficial non-specific effects (NSEs) following heterologous stimuli in humans and mice. The NSEs from BCG could be related to both adaptive and innate immune responses. The latter is also known as trained immunity (TI), a recently described biological feature of innate cells that enables functional improvement based on metabolic and epigenetic reprogramming. Currently, the mechanisms related to BCG-mediated TI are the focus of intense research, but many gaps are still in need of elucidation. This review discusses the present understanding of TI induced by BCG, exploring signaling pathways that are crucial to a trained phenotype in hematopoietic stem cells and monocytes/macrophages lineage. It focuses on BCG-mediated TI mechanisms, including the metabolic-epigenetic axis and the inflammasome pathway in these cells against intracellular pathogens. Moreover, this study explores the TI in different immune cell types, its ability to protect against various intracellular infections, and the integration of trained innate memory with adaptive memory to shape next-generation vaccines.
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Exosomes, organelles measuring 30-200nm, are secreted by various cell types. Leishmania exosomes consist of many proteins, including heat shock proteins, annexins, Glycoprotein 63, proteins exerting signaling activity and those containing mRNA and miRNA. Studies have demonstrated that Leishmania donovani exosomes downregulate IFN-γ and inhibit the expression of microbicidal molecules, such as TNF and nitric oxide, thus creating a microenvironment favoring parasite proliferation. Despite lacking immunological memory, data in the literature suggest that, following initial stimulation, mononuclear phagocytes may become "trained" to respond more effectively to subsequent stimuli. Here we characterized the effects of macrophage sensitization using L. braziliensis exosomes prior to infection by the same pathogen. Human macrophages were stimulated with L. braziliensis exosomes and then infected with L. braziliensis. Higher levels of IL-1ß and IL-6 were detected in cultures sensitized prior to infection compared to unstimulated infected cells. Moreover, stimulation with L. braziliensis exosomes induced macrophage production of IL-1ß, IL-6, IL-10 and TNF. Inhibition of exosome secretion by L. braziliensis prior to macrophage infection reduced cytokine production and produced lower infection rates than untreated infected cells. Exosome stimulation also induced the consumption/regulation of NLRP3 inflammasome components in macrophages, while the blockade of NLRP3 resulted in lower levels of IL-6 and IL-1ß. Our results suggest that L. braziliensis exosomes stimulate macrophages, leading to an exacerbated inflammatory state that may be NLRP3-dependent.
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Exossomos , Leishmania braziliensis , Leishmania donovani , Humanos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Interleucina-6/farmacologia , MacrófagosRESUMO
The bacillus Calmette-Guérin (BCG) can elicit enhanced innate immune responses against a wide range of infections, known as trained immunity. Brucella abortus is the causative agent of brucellosis, a debilitating disease that affects humans and animals. In this study, we demonstrate that C57BL/6 mouse bone marrow-derived macrophages under BCG training enhance inflammatory responses against B. abortus. BCG-trained macrophages showed increased MHC class II and CD40 expression on the cell surface and higher IL-6, IL-12, and IL-1ß production. The increase in IL-1ß secretion was accompanied by enhanced activation of canonical and noncanonical inflammasome platforms. We observed elevated caspase-11 expression and caspase-1 processing in BCG-trained macrophages in response to B. abortus compared with untrained cells. In addition, these BCG-trained cells showed higher NLRP3 expression after B. abortus infection. From a metabolic point of view, signaling through the Akt/mammalian target of rapamycin/S6 kinase pathway was also enhanced. In addition, BCG training resulted in higher inducible NO synthase expression and nitrite production, culminating in an improved macrophage-killing capacity against intracellular B. abortus. In vivo, we monitored a significant reduction in the bacterial burden in organs from BCG-trained C57BL/6 mice when compared with the untrained group. In addition, previous BCG immunization of RAG-1-deficient mice partially protects against Brucella infection, suggesting the important role of the innate immune compartment in this scenario. Furthermore, naive recipient mice that received BM transfer from BCG-trained donors showed greater resistance to B. abortus when compared with their untrained counterparts. These results demonstrate that BCG-induced trained immunity in mice results in better control of intracellular B. abortus in vivo and in vitro.
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Brucella abortus , Brucelose , Humanos , Animais , Camundongos , Vacina BCG , Camundongos Endogâmicos C57BL , Macrófagos , Brucelose/metabolismo , Caspases/metabolismo , MamíferosRESUMO
Despite the importance of the respiratory route for Brucella transmission, the lung immune response to this pathogen is scarcely characterized. We investigated the role of the cGAS/STING pathway of microbial DNA recognition in the control of respiratory Brucella infection. After in vitro B. abortus infection, CFU numbers were significantly higher in alveolar macrophages (AM) and lung explants from STING KO mice than in samples from wild type (WT) mice, but no difference was observed for cGAS KO samples. CFU were also increased in WT AM and lung epithelial cells preincubated with the STING inhibitor H151. Several proinflammatory cytokines (TNF-α, IL-1ß, IL-6, IP-10/CXCL10) were diminished in Brucella-infected lung explants and/or AM from STING KO mice and cGAS KO mice. These cytokines were also reduced in infected AM and lung epithelial cells pretreated with H151. After intratracheal infection with B. abortus, STING KO mice exhibited increased CFU in lungs, spleen and liver, a reduced expression of IFN-ß mRNA in lungs and spleen, and reduced levels of proinflammatory cytokines and chemokines in bronchoalveolar lavage fluid (BALF) and lung homogenates. Increased lung CFU and reduced BALF cytokines were also observed in cGAS KO mice. In summary, the cGAS/STING pathway induces the production of proinflammatory cytokines after respiratory Brucella infection, which may contribute to the STING-dependent control of airborne brucellosis.
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Brucelose Bovina , Brucelose , Animais , Camundongos , Bovinos , Brucella abortus , Citocinas/metabolismo , Nucleotidiltransferases/genéticaRESUMO
COVID-19 has accounted for more than 6 million deaths worldwide. Bacillus Calmette-Guérin (BCG), the existing tuberculosis vaccine, is known to induce heterologous effects over other infections due to trained immunity and has been proposed to be a potential strategy against SARS-CoV-2 infection. In this report, we constructed a recombinant BCG (rBCG) expressing domains of the SARS-CoV-2 nucleocapsid and spike proteins (termed rBCG-ChD6), recognized as major candidates for vaccine development. We investigated whether rBCG-ChD6 immunization followed by a boost with the recombinant nucleocapsid and spike chimera (rChimera), together with alum, provided protection against SARS-CoV-2 infection in K18-hACE2 mice. A single dose of rBCG-ChD6 boosted with rChimera associated with alum elicited the highest anti-Chimera total IgG and IgG2c Ab titers with neutralizing activity against SARS-CoV-2 Wuhan strain when compared with control groups. Importantly, following SARS-CoV-2 challenge, this vaccination regimen induced IFN-γ and IL-6 production in spleen cells and reduced viral load in the lungs. In addition, no viable virus was detected in mice immunized with rBCG-ChD6 boosted with rChimera, which was associated with decreased lung pathology when compared with BCG WT-rChimera/alum or rChimera/alum control groups. Overall, our study demonstrates the potential of a prime-boost immunization system based on an rBCG expressing a chimeric protein derived from SARS-CoV-2 to protect mice against viral challenge.
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COVID-19 , Mycobacterium bovis , Animais , Camundongos , Vacina BCG/genética , Proteínas Recombinantes de Fusão/genética , SARS-CoV-2 , Vacinas Sintéticas , COVID-19/prevenção & controle , Mycobacterium bovis/genéticaRESUMO
Nearly half of the natural disasters in the world are due to hydro-geomorphological hazards. Therefore, rainfall forecast is a key parameter for the implementation of landslides and flash-floods early warning systems. In this work we developed a routine in R software that enables the validation of a 3-day rainfall forecast by comparison with the daily rainfall data recorded in 101 automatic meteorological stations available in mainland Portugal. The routine integrates the pre-processing of base data, the matching between the 3-day rainfall forecast and the daily rainfall registered in the automatic meteorological stations based on sequence of days, the estimation of the difference between the forecasted and the real rainfall values and the computation of error measures, such as the bias, the mean absolute error, the mean absolute percentage error and the root mean square error. The results from the error measures, estimated for the 101 automatic meteorological stations, are then exported to an excel file. The routine is implemented for mainland Portugal and tested using data from February 2015, however, the spatial and temporal data can be easily updated for other regions.â¢A routine to validate the rainfall forecast at the regional scale using R programming language is implemented.â¢The automated routine can be easily updated and adapted with different spatial and temporal scales.
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Intracellular parasites from the Leishmania genus cause Leishmaniasis, a disease affecting millions of people worldwide. NLRP3 inflammasome is key for disease outcome, but the molecular mechanisms upstream of the inflammasome activation are still unclear. Here, we demonstrate that despite the absence of pyroptosis, Gasdermin-D (GSDMD) is active at the early stages of Leishmania infection in macrophages, allowing transient cell permeabilization, potassium efflux, and NLRP3 inflammasome activation. Further, GSDMD is processed into a non-canonical 25 kDa fragment. Gsdmd-/- macrophages and mice exhibit less NLRP3 inflammasome activation and are highly susceptible to infection by several Leishmania species, confirming the role of GSDMD for inflammasome-mediated host resistance. Active NLRP3 inflammasome and GSDMD are present in skin biopsies of patients, demonstrating activation of this pathway in human leishmaniasis. Altogether, our findings reveal that Leishmania subverts the normal functions of GSDMD, an important molecule to promote inflammasome activation and immunity in Leishmaniasis.
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Leishmania , Leishmaniose , Humanos , Camundongos , Animais , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Gasderminas , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Leishmania/metabolismo , Piroptose/fisiologiaRESUMO
Although the baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) infects lepidopteran invertebrates as natural hosts, represents an efficient vector for vaccine development. Baculovirus surface display induces strong humoral responses against viruses and parasites. A novel strategy based on capsid display carrying foreign antigens in the AcMNPV particle further improved the immune response by eliciting CD8+ T cell activation. In this study, we analyze the intracellular mechanisms and signalling pathways involved in CD8+ T cell activation by capsid display. Our results show that baculovirus can attach to the cell surface, enter dendritic cells (DCs), transit within endocytic vesicles and escape to the cytosol for further degradation by the proteasome. We found that the availability of viral proteins, endosomal acidification, and proteasome activity are needed for efficient Major Histocompatibility Complex class-I presentation by baculovirus carrying Ovalbumin in the viral capsid. Importantly, we demonstrated with this strategy that the induction of cytotoxic T cells and IL-12 production by DCs are TLR9-dependent and STING-independent. Finally, our study shows differential intracellular processing for capsid and surface baculovirus proteins in DCs and highlights the role of different danger receptors during cytotoxic T cell priming through the capsid display delivery system, which could lead to improved baculovirus-based vaccines development.
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Antineoplásicos , Baculoviridae , Baculoviridae/genética , Baculoviridae/metabolismo , Capsídeo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas do Capsídeo/genéticaRESUMO
Programmed cell death (PCD) is an important mechanism of innate immunity against bacterial pathogens. The innate immune PCD pathway involves the molecules caspase-7 and caspase-8, among others. Brucella abortus is a gram-negative bacterium that causes a zoonotic disease termed brucellosis. The innate immune response against this pathogen involves activation of inflammasome components and induction of pyroptosis. However, no studies so far have revealed the role of caspase-7 or caspase-8 during this bacterial infection. Herein, we demonstrate that caspase-7 is dispensable for caspase-1 processing, IL-1ß secretion and cell death in macrophages. Additionally, caspase-7 deficient animals control B. abortus infection as well as the wild type mice. Furthermore, we addressed the role of caspase-8 in inflammasome activation and pyroptosis during this bacterial infection. Macrophages deficient in caspase-8 secreted reduced amounts of IL-1ß that parallels with diminished caspase-1 activity when compared to wild type cells. Additionally, caspase-8 KO macrophages showed reduced LDH release when compared to wild type, suggesting that caspase-8 may play an important role in pyroptosis in response to B. abortus. Finally, caspase-8 KO animals were more susceptible to Brucella infection when compared to wild type mice. Overall, this study contributes to a better understanding of the involvement of caspase-7 and caspase-8 in innate immunity against B. abortus infection.
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Schistosomiasis is a chronic human parasitic disease that causes serious health problems worldwide. The disease-associated liver pathology is one of the hallmarks of infections by Schistosoma mansoni and Schistosoma japonicum, and is accountable for the debilitating condition found in infected patients. In the past few years, investigative studies have highlighted the key role played by neutrophils and the influence of inflammasome signalling pathway in different pathological conditions. However, it is noteworthy that the study of inflammasome activation in neutrophils has been overlooked by reports concerning macrophages and monocytes. This interplay between neutrophils and inflammasomes is much more poorly investigated during schistosomiasis. Herein, we reviewed the role of neutrophils during schistosomiasis and addressed the potential connection between these cells and inflammasome activation in this context.
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Hepatopatias , Schistosoma japonicum , Esquistossomose , Animais , Humanos , Inflamassomos/metabolismo , Neutrófilos/metabolismo , Schistosoma japonicum/fisiologia , Schistosoma mansoniRESUMO
Endoplasmic reticulum (ER) stress plays a major role in several inflammatory disorders. ER stress induces the unfolded protein response (UPR), a conserved response broadly associated with innate immunity and cell metabolic function in various scenarios. Brucella abortus, an intracellular pathogen, triggers the UPR via Stimulator of interferon genes (STING), an important regulator of macrophage metabolism during B. abortus infection. However, whether ER stress pathways underlie macrophage metabolic function during B. abortus infection remains to be elucidated. Here, we showed that the UPR sensor inositol-requiring enzyme 1α (IRE1α) is as an important component regulating macrophage immunometabolic function. In B. abortus infection, IRE1α supports the macrophage inflammatory profile, favoring M1-like macrophages. IRE1α drives the macrophage metabolic reprogramming in infected macrophages, contributing to the reduced oxidative phosphorylation and increased glycolysis. This metabolic reprogramming is probably associated with the IRE1α-dependent expression and stabilization of hypoxia-inducible factor-1 alpha (HIF-1α), an important molecule involved in cell metabolism that sustains the inflammatory profile in B. abortus-infected macrophages. Accordingly, we demonstrated that IRE1α favors the generation of mitochondrial reactive oxygen species (mROS) which has been described as an HIF-1α stabilizing factor. Furthermore, in infected macrophages, IRE1α drives the production of nitric oxide and the release of IL-1ß. Collectively, these data unravel a key mechanism linking the UPR and the immunometabolic regulation of macrophages in Brucella infection and highlight IRE1α as a central pathway regulating macrophage metabolic function during infectious diseases.
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Brucella abortus , Brucelose Bovina , Macrófagos , Animais , Bovinos , Brucella abortus/genética , Brucelose Bovina/metabolismo , Estresse do Retículo Endoplasmático/genética , Estresse do Retículo Endoplasmático/fisiologia , Endorribonucleases/genética , Endorribonucleases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismoRESUMO
PURPOSE: We present a physician survey of the impact of 21-gene Breast Recurrence Score test results on treatment decisions in clinical practice in Latin America. METHODS: This prospective survey enrolled consecutive patients at 14 sites in Argentina, Colombia, Mexico, and Peru who had routine 21-gene testing. Physician surveys captured patient and tumor characteristics and treatment decisions before and after 21-gene test results. The survey spanned the period before and after Trial Assigning Individualized Options for Treatment (TAILORx) results reported (June 2018). Overall net percent change in adjuvant chemotherapy recommendations was estimated, and asymptotic 95% CIs with continuity correction were calculated. The proportion with a change between pretest treatment recommendation and actual treatment received was calculated overall and by Recurrence Score groups per TAILORx. RESULTS: Between March 2015 and December 2019, the survey was completed for 647 patients; 20% were node-positive. The mean patient age was 54 years (24-85 years); 55% were postmenopausal; 17%, 63%, and 20% had grade 1, 2, and 3 tumors, respectively; and 30% had tumors > 2 cm. Recurrence Score (RS) results were as follows: 20% RS 0-10, 56% RS 11-25, and 24% RS 26-100. Overall, chemotherapy recommendations fell by a relative proportion of 39% (95% CI, 33.4 to 44.3) after 21-gene testing (33% decrease in node-negative and 55% decrease in node-positive). Among node-negative patients, the relative decrease in chemotherapy recommendations was 28% (95% CI, 18.9 to 39.5) before TAILORx and 36% (95% CI, 28.4 to 43.7) after. CONCLUSION: To our knowledge, this large survey of 21-gene test practice patterns was the first conducted in Latin America and showed the relevance of 21-gene testing in low- and medium-resource countries to minimize chemotherapy overuse and underuse in breast cancer. The results showed substantial reductions in chemotherapy use overall-especially after TAILORx reported-indicating the practice-changing potential of that study.
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Neoplasias da Mama , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Feminino , Perfilação da Expressão Gênica , Humanos , América Latina , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Estudos Prospectivos , Adulto JovemRESUMO
Bacillus Calmette-Guerin (BCG) is the only FDA approved first line therapy for patients with nonmuscle invasive bladder cancer. The purpose of this study is to better understand the role of innate immune pathways involved in BCG immunotherapy against murine bladder tumor. We first characterized the immunological profile induced by the MB49 mouse urothelial carcinoma cell line. MB49 cells were not able to activate an inflammatory response (TNF-α, IL-6, CXCL-10 or IFN-ß) after the stimulus with different agonists or BCG infection, unlike macrophages. Although MB49 cells are not able to induce an efficient immune response, BCG treatment could activate other cells in the tumor microenvironment (TME). We evaluated BCG intratumoral treatment in animals deficient for different innate immune molecules (STING-/-, cGAS-/-, TLR2-/-, TLR3-/-, TLR4-/-, TLR7-/-, TLR9-/-, TLR3/7/9-/-, MyD88-/-, IL-1R-/-, Caspase1/11-/-, Gasdermin-D-/- and IFNAR-/-) using the MB49 subcutaneous mouse model. Only MyD88-/- partially responded to BCG treatment compared to wild type (WT) mice, suggesting a role played by this adaptor molecule. Additionally, BCG intratumoral treatment regulates cellular infiltrate in TME with an increase of inflammatory macrophages, neutrophils and CD8+ T lymphocytes, suggesting an immune response activation that favors tumor remission in WT mice but not in MyD88-/-. The experiments using MB49 cells infected with BCG and co-cultured with macrophages also demonstrated that MyD88 is essential for an efficient immune response. Our data suggests that BCG immunotherapy depends partially on the MyD88-related innate immune pathway.
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Imunoterapia , Microambiente Tumoral , Neoplasias da Bexiga Urinária , Animais , Carcinoma de Células de Transição , Linhagem Celular Tumoral , Modelos Animais de Doenças , CamundongosRESUMO
In this study, we provide evidence that galectin-3 (Gal-3) plays an important role in Brucella abortus infection. Our results showed increased Gal-3 expression and secretion in B. abortus infected macrophages and mice. Additionally, our findings indicate that Gal-3 is dispensable for Brucella-containing vacuoles disruption, inflammasome activation and pyroptosis. On the other hand, we observed that Brucella-induced Gal-3 expression is crucial for induction of molecules associated to type I IFN signalling pathway, such as IFN-ß: Interferon beta (IFN-ß), C-X-C motif chemokine ligand 10 (CXCL10) and guanylate-binding proteins. Gal-3 KO macrophages showed reduced bacterial numbers compared to wild-type cells, suggesting that Gal-3 facilitates bacterial replication in vitro. Moreover, priming Gal-3 KO cells with IFN-ß favoured B. abortus survival in macrophages. Additionally, we also observed that Gal-3 KO mice are more resistant to B. abortus infection and these animals showed elevated production of proinflammatory cytokines when compared to control mice. Finally, we observed an increased recruitment of macrophages, dendritic cells and neutrophils in spleens of Gal-3 KO mice compared to wild-type animals. In conclusion, this study demonstrated that Brucella-induced Gal-3 is detrimental to host and this molecule is implicated in inhibition of recruitment and activation of immune cells, which promotes B. abortus spread and aggravates the infection. TAKE AWAYS: Brucella abortus infection upregulates galectin-3 expression Galectin-3 regulates guanylate-binding proteins expression but is not required for Brucella-containing vacuole disruption Galectin-3 modulates proinflammatory cytokine production during bacterial infection Galectin-3 favours Brucella replication.