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1.
Infect Drug Resist ; 14: 4409-4419, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34729017

RESUMO

Background: Human listeriosis is a severe foodborne infection caused by Listeria monocytogenes and the data of patients with this infection are largely limited for the Hefei population. Purpose: This is a retrospective study that evaluated the clinical and laboratory data of patients with listeriosis at a tertiary hospital in Hefei City. Patients and Methods: A total of 24 listeriosis patients were admitted to the First Affiliated Hospital of Anhui Medical University from January 2003 to July 2021. Data from all patients were collected from the hospital's electronic medical records. Results: The most common symptom of all patients was fever (91.7%), followed by altered consciousness (50.0%), rashes (45.8%), respiratory distress symptoms (37.5%), nuchal rigidity (29.2%), and headaches (20.8%). Laboratory results also indicated elevated C-reactive protein (CRP) (79.1%), hypoproteinemia (75.0%), anemia (62.5%), leukocytosis (45.8%), and neutrophilia (45.8%). The mean value of 5.1 µg/mL (SD, 3.8) for D-dimer (D-D) was significantly higher than the normal value ((0.00-0.50) µg/mL), while both altered consciousness (6 vs 4, P = 0.034) and headaches (4 vs 1, P = 0.036) occurred more frequently in the neurolisteriosis group compared with the bacteremia one. Additionally, the mean maximal body temperature (°C) (40.5 ± 0.7) as well as white blood cell (WBC) (15.3 vs 7.5 ×109/L, P = 0.014) and neutrophil (NEUT#) (13.2 vs 6.1 ×109/L, P = 0.026) counts of neurolisteriosis patients were higher than those of bacteremia (39.4 ± 0.4) (P = 0.001). Of all patients, four (50%) from the maternal-neonatal group remained uncured. Conclusion: Listeriosis is a rare disease with extremely variable clinical characteristics in Hefei City. Our data indicated that unexplained fever, altered consciousness, hypoproteinemia, anemia, elevated CRP and DD should be considered to assist diagnosis of listeriosis for early treatment interventions.

2.
Biochem Soc Trans ; 2021 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-34812891

RESUMO

Pyroptosis is a highly inflammatory and lytic type of programmed cell death (PCD) commenced by inflammasomes, which sense perturbations in the cytosolic environment. Recently, several ground-breaking studies have linked a family of pore-forming proteins known as gasdermins (GSDMs) to pyroptosis. The human genome encodes six GSDM proteins which have a characteristic feature of forming pores in the plasma membrane resulting in the disruption of cellular homeostasis and subsequent induction of cell death. GSDMs have an N-terminal cytotoxic domain and an auto-inhibitory C-terminal domain linked together through a flexible hinge region whose proteolytic cleavage by various enzymes releases the N-terminal fragment that can insert itself into the inner leaflet of the plasma membrane by binding to acidic lipids leading to pore formation. Emerging studies have disclosed the involvement of GSDMs in various modalities of PCD highlighting their role in diverse cellular and pathological processes. Recently, the cryo-EM structures of the GSDMA3 and GSDMD pores were resolved which have provided valuable insights into the pore formation process of GSDMs. Here, we discuss the current knowledge regarding the role of GSDMs in PCD, structural and molecular aspects of autoinhibition, and pore formation mechanism followed by a summary of functional consequences of gasdermin-induced membrane permeabilization.

3.
Anal Chem ; 93(42): 14238-14246, 2021 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-34636246

RESUMO

Direct detection of SARS-CoV-2 in biological specimens is often challenging due to the low abundance of viral components and lack of enough sensitivity. Herein, we developed a new type of chemiluminescent functionalized magnetic nanomaterial for sensitive detection of the SARS-CoV-2 antigen. First, HAuCl4 was reduced by N-(aminobutyl)-N-(ethylisoluminol) (ABEI) in the presence of amino magnetic beads (MB-NH2) to generate ABEI-AuNPs, which were directly assembled on the surface of MB-NH2. Then, Co2+ was modified onto the surface to form MB@ABEI-Au/Co2+ (MAA/Co2+). MAA/Co2+ exhibited good chemiluminescence (CL) and magnetic properties. It was also found that it was easy for the antibody to be connected with MAA/Co2+. Accordingly, MAA/Co2+ was used as a sensing interface to construct a label-free immunoassay for rapid detection of the N protein in SARS-CoV-2. The immunoassay showed a linear range from 0.1 pg/mL to 10 ng/mL and a low detection limit of 69 fg/mL, which was superior to previously reported methods for N protein detection. It also demonstrated good selectivity by virtue of magnetic separation, which effectively removed a sample matrix after immunoreactions. It was successfully applied for the detection of the N protein in spiked human serum and saliva samples. Furthermore, the immunoassay was integrated with an automatic CL analyzer with magnetic separation to detect the N protein in patient serums and rehabilitation patient serums with satisfactory results. Thus, the CL immunoassay without a complicated labeling procedure is sensitive, selective, fast, simple, and cost-effective, which may be used to combat the COVID-19 pandemic. Finally, the CL quenching mechanism of the N protein in the immunoassay was also explored.


Assuntos
COVID-19 , Nanopartículas Metálicas , Ouro , Humanos , Imunoensaio , Limite de Detecção , Luminescência , Medições Luminescentes , Pandemias , SARS-CoV-2
4.
Sens Actuators B Chem ; 349: 130739, 2021 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-34611381

RESUMO

Accurate and efficient early diagnosis is crucial for the control of COVID-19 pandemic. However, methods that can balance sensitivity, high throughput, detection speed and automation simultaneously are still scarce. Here, we report an automatic label-free chemiluminescence immunoassay (CLIA) for rapid SARS-CoV-2 nucleocapsid protein (NP) detection with high sensitivity and throughput. N-(4-aminobutyl)-N-ethylisoluminol and Co2+ dual-functionalized chemiluminescent magnetic beads (dfCL-MB) were first applied to the detection of protein by a novel and simple strategy. Sulphydryl polyethylene glycol was coated on the surface of dfCL-MB so as to assemble dfCL-MB and antibody conjugated gold nanoparticles through Au-S bond. Considering the high-risk application scenarios, the immunosensor was integrated with an automatic chemiluminescence analyzer so that the whole testing procedure could be carried out automatically without manual operation. A linear correlation between CL intensities and the logarithm of NP concentration was obtained in the range of 0.1-10,000 pg/mL with a detection limit of 21 fg/mL. The whole process cost 25 min and the sample compartment can bear 24 samples simultaneously. The spiked human serum samples and serum samples from COVID-19 patients were determined with satisfactory recoveries of 91.1-109.4%, suggesting that the proposed label-free CLIA is of great potential for SARS-CoV-2 NP detection in practice.

5.
Expert Rev Vaccines ; : 1-13, 2021 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-34633259

RESUMO

INTRODUCTION: Vaccines are the agreed upon weapon against the COVID-19 pandemic. This review discusses about COVID-19 subunit vaccines adjuvants and their signaling pathways, which could provide a glimpse into the selection of appropriate adjuvants for prospective vaccine development studies.Areas covered: In the introduction, a brief background about the SARS-CoV-2 pandemic, the vaccine development race and classes of vaccine adjuvants were provided. . The antigen, trial stage, and types of adjuvants were extracted from the included articles and thun assimilated. Finally, the pattern recognition receptors (PRRs), their classes, cognate adjuvants, and potential signaling pathways were comprehended.Expert opinion: Adjuvants are unsung heroes of subunit vaccines. The in silico studies are very vital in avoiding several costly trial errors and save much work times. The majority of the (pre)clinical studies are promising. It is encouraging that most of the selected adjuvants are novel. Much emphasis must be paid to the optimal paring of antigen-adjuvant-PRRs for obtaining the desired vaccine effect. A good subunit vaccine/adjuvant is one that has high efficacy, safety, dose sparing, and rapid seroconversion rate and broad spectrum of immune response. In the years to come, COVID-19 adjuvanted subunit vaccines are expected to have superior utility than any other vaccines for various reasons.

6.
Biochem Biophys Res Commun ; 581: 38-45, 2021 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-34653677

RESUMO

A detrimental role of the receptor for the advanced glycation end product (RAGE) has been identified in the immune response, and various pathological conditions and its V and C1 domains in the extracellular region of RAGE are believed to be the main ligand-binding domains. Consequently, specific inhibitors targeting those domains could be of clinical value in fighting against the pathological condition associated with RAGE over-activation. Single-domain antibodies, also called nanobodies (Nbs), are antibody fragments engineered from the heavy-chain only antibodies found in camelids, which offer a range of advantages in therapy. In this study, we report the development and characterization of the V-C1 domain-specific Nbs. Three Nbs (3CNB, 4BNB, and 5ENB) targeting V-C1 domain of human RAGE were isolated from an immunized alpaca using a phage display. All of these Nbs revealed high thermostability. 3CNB, 4BNB, and 5ENB bind to V-C1 domain with a dissociation constant (KD) of 27.25, 39.37, and 47.85 nM, respectively, using Isothermal Titration Calorimetry (ITC). After homodimerization using human IgG1-Fc fusion, their binding affinity improved to 0.55, 0.62, and 0.41 nM, respectively, using Surface Plasmon Resonance (SPR). Flow cytometry showed all the Fc fusions Nbs can bind to human RAGE expressed on the cell surface. Competitive ELISA further confirmed their V-C1-hS100B blocking ability in solution, providing insights into the applicability of Nbs in treating RAGE-associated diseases.

7.
Cell Rep ; 36(11): 109708, 2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34506741

RESUMO

Cellular immunity is important in determining the disease severity of COVID-19 patients. However, current understanding of SARS-CoV-2 epitopes mediating cellular immunity is limited. Here we apply T-Scan, a recently developed method, to identify CD8+ T cell epitopes from COVID-19 patients of four major HLA-A alleles. Several identified epitopes are conserved across human coronaviruses, which might mediate pre-existing cellular immunity to SARS-CoV-2. In addition, we identify and validate four epitopes that were mutated in the newly circulating variants, including the Delta variant. The mutations significantly reduce T cell responses to the epitope peptides in convalescent and vaccinated samples. We further determine the crystal structure of HLA-A∗02:01/HLA-A∗24:02 in complex with the epitope KIA_S/NYN_S, respectively, which reveals the importance of K417 and L452 of the spike protein for binding to HLA. Our data suggest that evading cellular immunity might contribute to the increased transmissibility and disease severity associated with the new SARS-CoV-2 variants.


Assuntos
Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Epitopos de Linfócito T/imunologia , Imunidade Celular/imunologia , SARS-CoV-2/imunologia , Sequência de Aminoácidos , Humanos , Glicoproteína da Espícula de Coronavírus/imunologia
8.
Front Immunol ; 12: 715464, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34539645

RESUMO

The mutants resulted from the ongoing SARS-CoV-2 epidemic have showed resistance to antibody neutralization and vaccine-induced immune response. The present study isolated and identified two novel SARS-CoV-2 neutralizing antibodies (nAbs) from convalescent COVID-19 patients. These two nAbs (XG81 and XG83) were then systemically compared with nine nAbs that were reconstructed by using published data, and revealed that, even though these two nAbs shared targeting epitopes on spike protein, they were different from any of the nine nAbs. Compared with XG81, XG83 exhibited a higher RBD binding affinity and neutralization potency against wild-typed pseudovirus, variant pseudoviruses with mutated spike proteins, such as D614G, E484Q, and A475V, as well as the authentic SARS-CoV-2 virus. To explore potential broadly neutralizing antibodies, heavy and light chains from all 18 nAbs (16 published nAbs, XG81 and XG83) were cross-recombined, and some of the functional antibodies were screened and studied for RBD binding affinity, and neutralizing activity against pseudovirus and the authentic SARS-CoV-2 virus. The results demonstrated that several recombined antibodies had a more potent neutralization activity against variant pseudoviruses compared with the originally paired Abs. Taken together, the novel neutralizing antibodies identified in this study are a likely valuable addition to candidate antibody drugs for the development of clinical therapeutic agents against SARS-CoV-2 to minimize mutational escape.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Anticorpos Amplamente Neutralizantes/imunologia , Anticorpos Amplamente Neutralizantes/uso terapêutico , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/genética , Anticorpos Antivirais/uso terapêutico , Afinidade de Anticorpos/imunologia , Linfócitos B/imunologia , Anticorpos Amplamente Neutralizantes/genética , COVID-19/imunologia , COVID-19/terapia , Linhagem Celular , Epitopos/imunologia , Humanos , Imunoterapia/métodos , Testes de Neutralização , SARS-CoV-2/efeitos dos fármacos
9.
Pathogens ; 10(7)2021 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-34358031

RESUMO

The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global concern. Immunoglobin A (IgA) contributes to virus neutralization at the early stage of infection. Longitudinal studies are needed to assess whether SARS-CoV-2-specific IgA production persists for a longer time in patients recovered from severe COVID-19 and its lasting symptoms that can have disabling consequences should also be alerted to susceptible hosts. Here, we tracked the anti-SARS-CoV-2 spike protein receptor-binding domain (RBD) antibody levels in a cohort of 88 COVID-19 patients. We found that 52.3% of the patients produced more anti-SARS-CoV-2 RBD IgA than IgG or IgM, and the levels of IgA remained stable during 4-41 days of infection. One of these IgA-dominant COVID-19 patients, concurrently with IgA nephropathy (IgAN), presented with elevated serum creatinine and worse proteinuria during the infection, which continued until seven months post-infection. The serum levels of anti-SARS-CoV-2 RBD and total IgA were higher in this patient than in healthy controls. Changes in the composition of the intestinal microbiota, increased IgA highly coated bacteria, and elevated concentration of the proinflammatory cytokine IL-18 were indicative of potential involvement of intestinal dysbiosis and inflammation to the systemic IgA level and, consequently, the disease progression. Collectively, our work highlighted the potential adverse effect of the mucosal immune response to SARS-CoV-2 infection, and that additional care should be taken with COVID-19 patients presenting with chronic diseases such as IgAN.

10.
J Exp Med ; 218(9)2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34342641

RESUMO

Activation of NLRP3 inflammasome is precisely controlled to avoid excessive activation. Although multiple molecules regulating NLRP3 inflammasome activation have been revealed, the checkpoints governing NLRP3 inflammasome activation remain elusive. Here, we show that activation of NLRP3 inflammasome is governed by GSTO1-promoted ASC deglutathionylation in macrophages. Glutathionylation of ASC inhibits ASC oligomerization and thus represses activation of NLRP3 inflammasome in macrophages, unless GSTO1 binds ASC and deglutathionylates ASC at ER, under control of mitochondrial ROS and triacylglyceride synthesis. In macrophages expressing ASCC171A, a mutant ASC without glutathionylation site, activation of NLRP3 inflammasome is GSTO1 independent, ROS independent, and signal 2 less dependent. Moreover, AscC171A mice exhibit NLRP3-dependent hyperinflammation in vivo. Our results demonstrate that glutathionylation of ASC represses NLRP3 inflammasome activation, and GSTO1-promoted ASC deglutathionylation at ER, under metabolic control, is a checkpoint for activating NLRP3 inflammasome.

11.
ACS Sens ; 6(7): 2709-2719, 2021 07 23.
Artigo em Inglês | MEDLINE | ID: mdl-34263598

RESUMO

The spread of Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2), resulting in a global pandemic with around four million deaths. Although there are a variety of nucleic acid-based tests for detecting SARS-CoV-2, these methods have a relatively high cost and require expensive supporting equipment. To overcome these limitations and improve the efficiency of SARS-CoV-2 diagnosis, we developed a microfluidic platform that collected serum by a pulling-force spinning top and paper-based microfluidic enzyme-linked immunosorbent assay (ELISA) for quantitative IgA/IgM/IgG measurements in an instrument-free way. We further validated the paper-based microfluidic ELISA analysis of SARS-CoV-2 receptor-binding domain (RBD)-specific IgA/IgM/IgG antibodies from human blood samples as a good measurement with higher sensitivity compared with traditional IgM/IgG detection (99.7% vs 95.6%) for early illness onset patients. In conclusion, we provide an alternative solution for the diagnosis of SARS-CoV-2 in a portable manner by this smart integration of pulling-force spinning top and paper-based microfluidic immunoassay.


Assuntos
Teste para COVID-19 , COVID-19 , Ensaio de Imunoadsorção Enzimática , Dispositivos Lab-On-A-Chip , Anticorpos Antivirais/sangue , COVID-19/diagnóstico , Humanos , SARS-CoV-2 , Sensibilidade e Especificidade
12.
Infect Drug Resist ; 14: 2741-2752, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34295167

RESUMO

Background: Brucellosis is currently one of the most widespread zoonotic diseases caused by Brucella genus, and the Brucella melitensis is the major pathogen. The number of people infected with Brucella has gradually increased in Anhui Province. Purpose: To retrospectively evaluate the epidemiological, clinical, and laboratory data of brucellosis patients in Anhui Province. Patients and Methods: A total of 109 brucellosis patients were admitted to the First Affiliated Hospital of Anhui Medical University from January 2012 to March 2021. Data from all patients were retrieved from the hospital's electronic medical system. The final results were grouped and compared according to the presence or absence of bacteremic brucellosis and three phases of brucellosis. Results: The most common symptoms among all 109 brucellosis patients were fever (89.0%), followed by chills (52.3%), arthralgia (48.6%), and weight loss (30.3%), and laboratory results presented with anemia (65.1%), elevate of C-reactive protein (CRP) (91.7%), erythrocyte sedimentation rate (ESR) (86.2%), aspartate aminotransferase (AST) (40.4%), and lactate dehydrogenase (LDH) (43.1%). The percentage of fever (96.1%), arthralgia (58.8%), anorexia (35.3%), leukopenia (31.4%), and the AST (51.0%) were higher in bacteremic than nonbacteremic group. Additionally, the median level of LDH (332.0 mg/L, IQR, 209.0-553.0) was higher in bacteremic than nonbacteremic group. Nevertheless, the albumin (36.0 mg/L, IQR, 33.9-38.2) was lower in the bacteremic group. The percentage of fever (94.9%) and the median LDH level (316.0 U/L (IQR,218.0-517.5)) in the acute phase of brucellosis were higher than the percentage of fever (72.0%) and the median LDH level (209.0 U/L (IQR,162.0-276.0)) in the subacute phase of brucellosis. Conclusion: Brucellosis has become an important public health issue in Anhui Province. Brucellosis is a disease with diverse clinical manifestations. Our data showed that unexplained fever, arthralgia, and elevated AST and LDH should be considered as a diagnosis of bacteremia brucellosis for early treatment intervention.

13.
J Agric Food Chem ; 69(45): 13315-13322, 2021 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-34076413

RESUMO

Buckwheat is one of the five main allergenic foods (eggs, milk, wheat, buckwheat, and peanuts). Oleosin is an important type of allergen in some allergic foods. However, although most diagnostic nut and seed extracts are defatted, some patients with food allergies may have false negative diagnostic results of oleosin in vitro. Recently, we found that the serum of buckwheat allergic patients responded strongly to an 18 kDa protein. Mass spectrometry analysis showed it is the oleosin protein family. We further purified and evaluated the allergenicity of this buckwheat oleosin-type allergen, which is involved in the formation of buckwheat oil bodies. The tartary buckwheat oleosin allergen was named Fag t 6, according to the WHO/IUIS Allergen Nomenclature Subcommittee criteria. The DNA sequence of tartary buckwheat oleosin was cloned. Dot blot and enzyme-linked immunosorbent assay (ELISA) showed half of the 20 buckwheat allergic patients' serum had strong reactivity with purified buckwheat Fag t 6. Circular dichroism experiment analysis of its thermal stability showed a Tm of 64.65 ± 0.65 °C. A buckwheat allergy showed possible cross-reaction with a wheat allergy. In summary, this study not only increases our understanding of buckwheat allergies and oil-soluble allergens in general, it may also be used to improve diagnostic tests for buckwheat allergies in the future.


Assuntos
Fagopyrum , Hipersensibilidade Alimentar , Hipersensibilidade a Trigo , Alérgenos , Humanos , Proteínas de Plantas/genética , Sementes
14.
Front Mol Biosci ; 8: 670815, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34136533

RESUMO

The 2019-2020 winter was marked by the emergence of a new coronavirus (SARS-CoV-2) related disease (COVID-19), which started in Wuhan, China. Its high human-to-human transmission ability led to a worldwide spread within few weeks and has caused substantial human loss. Mechanical antiviral control approach, drug repositioning, and use of COVID-19 convalescent plasmas (CPs) were the first line strategies utilized to mitigate the viral spread, yet insufficient. The urgent need to contain this deadly pandemic has led searchers and pharmaceutical companies to develop vaccines. However, not all vaccines manufactured are safe. Besides, an alternative and effective treatment option for such an infectious disease would include pure anti-viral neutralizing monoclonal antibodies (NmAbs), which can block the virus at specific molecular targets from entering cells by inhibiting virus-cell structural complex formation, with more safety and efficiency than the CP. Indeed, there is a lot of molecular evidence about the protector effect and the use of molecular feature-based NmAbs as promising therapeutics to contain COVID-19. Thus, from the scientific publication database screening, we here retrieved antibody-related papers and summarized the repertory of characterized NmAbs against SARS-CoV-2, their molecular neutralization mechanisms, and their immunotherapeutic pros and cons. About 500 anti-SARS-CoV-2 NmAbs, characterized through competitive binding assays and neutralization efficacy, were reported at the writing time (January 2021). All NmAbs bind respectively to SARS-CoV-2 S and exhibit high molecular neutralizing effects against wild-type and/or pseudotyped virus. Overall, we defined six NmAb groups blocking SARS-CoV-2 through different molecular neutralization mechanisms, from which five potential neutralization sites on SARS-CoV-2 S protein are described. Therefore, more efforts are needed to develop NmAbs-based cocktails to mitigate COVID-19.

15.
Front Immunol ; 12: 636225, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33833757

RESUMO

Gain-of-function mutations in STING1 cause the monogenic interferonopathy, SAVI, which presents with early-onset systemic inflammation, cold-induced vasculopathy and/or interstitial lung disease. We identified 5 patients (3 kindreds) with predominantly peripheral vascular disease who harbor 3 novel STING1 variants, p.H72N, p.F153V, and p.G158A. The latter two were predicted by a previous cryo-EM structure model to cause STING autoactivation. The p.H72N variant in exon 3, however, is the first SAVI-causing variant in the transmembrane linker region. Mutations of p.H72 into either charged residues or hydrophobic residues all led to dramatic loss of cGAMP response, while amino acid changes to residues with polar side chains were able to maintain the wild type status. Structural modeling of these novel mutations suggests a reconciled model of STING activation, which indicates that STING dimers can oligomerize in both open and closed states which would obliviate a high-energy 180° rotation of the ligand-binding head for STING activation, thus refining existing models of STING activation. Quantitative comparison showed that an overall lower autoactivating potential of the disease-causing mutations was associated with less severe lung disease, more severe peripheral vascular disease and the absence of a robust interferon signature in whole blood. Our findings are important in understanding genotype-phenotype correlation, designing targeted STING inhibitors and in dissecting differentially activated pathways downstream of different STING mutations.


Assuntos
Inflamação/genética , Doenças Pulmonares Intersticiais/genética , Proteínas de Membrana/genética , Mutação , Doenças Vasculares Periféricas/genética , Adulto , Criança , Análise Mutacional de DNA , Feminino , Predisposição Genética para Doença , Células HEK293 , Humanos , Inflamação/diagnóstico , Inflamação/metabolismo , Inflamação/terapia , Doenças Pulmonares Intersticiais/diagnóstico , Doenças Pulmonares Intersticiais/metabolismo , Doenças Pulmonares Intersticiais/terapia , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Modelos Moleculares , Doenças Vasculares Periféricas/diagnóstico , Doenças Vasculares Periféricas/metabolismo , Doenças Vasculares Periféricas/terapia , Fenótipo , Conformação Proteica , Multimerização Proteica , Índice de Gravidade de Doença , Relação Estrutura-Atividade , Sequenciamento Completo do Exoma , Adulto Jovem
16.
Front Chem ; 9: 622898, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33889562

RESUMO

The Coronavirus disease-19 (COVID-19) pandemic is still devastating the world causing significant social, economic, and political chaos. Corresponding to the absence of globally approved antiviral drugs for treatment and vaccines for controlling the pandemic, the number of cases and/or mortalities are still rising. Current patient management relies on supportive treatment and the use of repurposed drugs as an indispensable option. Of a crucial role in the viral life cycle, ongoing studies are looking for potential inhibitors to the main protease (Mpro) of severe acute respiratory syndrome Coronavirus -2 (SARS-CoV-2) to tackle the pandemic. Although promising results have been achieved in searching for drugs inhibiting the Mpro, work remains to be done on designing structure-based improved drugs. This review discusses the structural basis of potential inhibitors targeting SARS-CoV-2 Mpro, identifies gaps, and provides future directions. Further, compounds with potential Mpro based antiviral activity are highlighted.

17.
J Cancer ; 12(8): 2295-2316, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33758607

RESUMO

Ovarian clear cell carcinoma (OCCC) is a special pathological type of epithelial ovarian carcinoma (EOC) and has a high prevalence in Asia without specific molecular subtype classification. Endometriosis is a recognized precancerous lesion that carries 3-fold increased risk of OCCC. Ovarian endometrioid carcinoma, which also originates from endometriosis, shares several features with OCCC, including platinum resistance and younger age at diagnosis. Patients with OCCC have about a 2.5 to 4 times greater risk of having a venous thromboembolism (VTE) compared with other EOC, and OCCC tends to metastasize through lymphatic vesicular and peritoneal spread as opposed to hematogenous metastasis. There is only mild elevation of the conventional biomarker CA125. Staging surgery or optimal cytoreduction combined with chemotherapy is a common therapeutic strategy for OCCC. However, platinum resistance commonly portends a poor prognosis, so novel treatments are urgently needed. Targeted therapy and immunotherapy are currently being studied, including PARP, EZH2, and ATR inhibitors combined with the synthetic lethality of ARID1A-dificiency, and MAPK/PI3K/HER2, VEGF/bFGF/PDGF, HNF1ß, and PD-1/PD-L1 inhibitors. Advanced stage, suboptimal cytoreduction, platinum resistance, lymph node metastasis, and VTE are major prognostic predictors for OCCC. We focus on update pathogenesis, diagnostic methods and therapeutic approaches to provide future directions for clinical diagnosis and treatment of OCCC.

18.
J Virol ; 2021 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-33658349

RESUMO

Cell entry by SARS-CoV-2 requires the binding between the receptor-binding domain (RBD) of the viral Spike protein and the cellular angiotensin-converting enzyme 2 (ACE2). As such, RBD has become the major target for vaccine development, while RBD-specific antibodies are pursued as therapeutics. Here, we report the development and characterization of SARS-CoV-2 RBD-specific VHH/nanobody (Nb) from immunized alpacas. Seven RBD-specific Nbs with high stability were identified using phage display. They bind to SARS-CoV-2 RBD with affinity KD ranging from 2.6 to 113 nM, and six of them can block RBD-ACE2 interaction. The fusion of the Nbs with IgG1 Fc resulted in homodimers with greatly improved RBD-binding affinities (KD ranging from 72.7 pM to 4.5 nM) and nanomolar RBD-ACE2 blocking abilities. Furthermore, the fusion of two Nbs with non-overlapping epitopes resulted in hetero-bivalent Nbs, namely aRBD-2-5 and aRBD-2-7, with significantly higher RBD binding affinities (KD of 59.2 pM and 0.25 nM) and greatly enhanced SARS-CoV-2 neutralizing potency. The 50% neutralization dose (ND50) of aRBD-2-5 and aRBD-2-7 was 1.22 ng/mL (∼0.043 nM) and 3.18 ng/mL (∼0.111 nM), respectively. These high-affinity SARS-CoV-2 blocking Nbs could be further developed into therapeutics as well as diagnostic reagents for COVID-19.ImportanceTo date, SARS-CoV-2 has caused tremendous loss of human life and economic output worldwide. Although a few COVID-19 vaccines have been approved in several countries, the development of effective therapeutics, including SARS-CoV-2 targeting antibodies, remains critical. Due to their small size (13-15 kDa), high solubility, and stability, Nbs are particularly well suited for pulmonary delivery and more amenable to engineer into multivalent formats than the conventional antibody. Here, we report a series of new anti-SARS-CoV-2 Nbs isolated from immunized alpaca and two engineered hetero-bivalent Nbs. These potent neutralizing Nbs showed promise as potential therapeutics against COVID-19.

19.
Toxins (Basel) ; 13(2)2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33572271

RESUMO

With the rapid growth of antibiotic-resistant bacteria, it is urgent to develop alternative therapeutic strategies. Pore-forming toxins (PFTs) belong to the largest family of virulence factors of many pathogenic bacteria and constitute the most characterized classes of pore-forming proteins (PFPs). Recent studies revealed the structural basis of several PFTs, both as soluble monomers, and transmembrane oligomers. Upon interacting with host cells, the soluble monomer of bacterial PFTs assembles into transmembrane oligomeric complexes that insert into membranes and affect target cell-membrane permeability, leading to diverse cellular responses and outcomes. Herein we have reviewed the structural basis of pore formation and interaction of PFTs with the host cell membrane, which could add valuable contributions in comprehensive understanding of PFTs and searching for novel therapeutic strategies targeting PFTs and interaction with host receptors in the fight of bacterial antibiotic-resistance.


Assuntos
Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Membrana Celular/metabolismo , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Animais , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/patogenicidade , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/química , Toxinas Bacterianas/antagonistas & inibidores , Toxinas Bacterianas/química , Membrana Celular/efeitos dos fármacos , Membrana Celular/microbiologia , Resistência a Medicamentos/efeitos dos fármacos , Interações Hospedeiro-Patógeno , Humanos , Terapia de Alvo Molecular , Proteínas Citotóxicas Formadoras de Poros/antagonistas & inibidores , Proteínas Citotóxicas Formadoras de Poros/química , Conformação Proteica , Relação Estrutura-Atividade , Virulência
20.
Nucleic Acids Res ; 49(5): 2959-2972, 2021 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-33619523

RESUMO

The interferon gamma-inducible protein 16 (IFI16) and its murine homologous protein p204 function in non-sequence specific dsDNA sensing; however, the exact dsDNA recognition mechanisms of IFI16/p204, which harbour two HIN domains, remain unclear. In the present study, we determined crystal structures of p204 HINa and HINb domains, which are highly similar to those of other PYHIN family proteins. Moreover, we obtained the crystal structure of p204 HINab domain in complex with dsDNA and provided insights into the dsDNA binding mode. p204 HINab binds dsDNA mainly through α2 helix of HINa and HINb, and the linker between them, revealing a similar HIN:DNA binding mode. Both HINa and HINb are vital for HINab recognition of dsDNA, as confirmed by fluorescence polarization assays. Furthermore, a HINa dimerization interface was observed in structures of p204 HINa and HINab:dsDNA complex, which is involved in binding dsDNA. The linker between HINa and HINb reveals dynamic flexibility in solution and changes its direction at ∼90° angle in comparison with crystal structure of HINab:dsDNA complex. These structural information provide insights into the mechanism of DNA recognition by different HIN domains, and shed light on the unique roles of two HIN domains in activating the IFI16/p204 signaling pathway.


Assuntos
DNA/química , Proteínas Nucleares/química , Fosfoproteínas/química , Cristalografia por Raios X , DNA/metabolismo , Modelos Moleculares , Proteínas Nucleares/metabolismo , Fosfoproteínas/metabolismo , Ligação Proteica , Domínios Proteicos , Multimerização Proteica
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