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1.
MAbs ; 14(1): 2020203, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35133949

RESUMO

Despite recent advances in transgenic animal models and display technologies, humanization of mouse sequences remains one of the main routes for therapeutic antibody development. Traditionally, humanization is manual, laborious, and requires expert knowledge. Although automation efforts are advancing, existing methods are either demonstrated on a small scale or are entirely proprietary. To predict the immunogenicity risk, the human-likeness of sequences can be evaluated using existing humanness scores, but these lack diversity, granularity or interpretability. Meanwhile, immune repertoire sequencing has generated rich antibody libraries such as the Observed Antibody Space (OAS) that offer augmented diversity not yet exploited for antibody engineering. Here we present BioPhi, an open-source platform featuring novel methods for humanization (Sapiens) and humanness evaluation (OASis). Sapiens is a deep learning humanization method trained on the OAS using language modeling. Based on an in silico humanization benchmark of 177 antibodies, Sapiens produced sequences at scale while achieving results comparable to that of human experts. OASis is a granular, interpretable and diverse humanness score based on 9-mer peptide search in the OAS. OASis separated human and non-human sequences with high accuracy, and correlated with clinical immunogenicity. BioPhi thus offers an antibody design interface with automated methods that capture the richness of natural antibody repertoires to produce therapeutics with desired properties and accelerate antibody discovery campaigns. The BioPhi platform is accessible at https://biophi.dichlab.org and https://github.com/Merck/BioPhi.


Assuntos
Aprendizado Profundo , Animais , Anticorpos , Camundongos
2.
Anal Chim Acta ; 1221: 340135, 2022 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-35934406

RESUMO

In recent years, some studies have found that oriented immobilization of antibodies to microspheres can fully expose the antigen binding sites of antibodies, which can improve the sensitivity of sandwich immunoassays for the detection of proteins. Can this antibody immobilization strategy also improve the sensitivity of competitive immunoassays for the detection of small molecules? To answer this question, the conjugate MS-SPG-Ab (oriented immobilization of aflatoxin B1 antibody to time-resolved fluorescent microspheres via streptococcal protein G) and the conjugate MS-Ab (nonoriented immobilization of aflatoxin B1 antibody to time-resolved fluorescent microspheres) were prepared, and a lateral flow immunoassay (LFIA) for the detection of aflatoxin B1 (AFB1) was established. The detection performance of the two methods was compared. The results showed that under the condition that the number of "effective" antibodies immobilized on TRF-MS was similar, compared with the nonoriented immobilization strategy (IC50 = 0.21 ng mL-1), the LFIA method established by the oriented immobilization strategy reduced the sensitivity of AFB1 detection (IC50 = 0.37 ng mL-1). However, this method can obtain higher detection precision for AFB1, the CV values were all below 8%. And it has stronger tolerance to the matrix of maize and peanut samples. The bias of LFIAs based on oriented immobilization technology (-14.93%-7.92%) was lower than nonoriented immobilization technology (28.16%-34.19%) for AFB1 detection in the two sample extracts. This study suggests that the LFIA method based on the oriented immobilization of antibodies can improve the accuracy of the detection results when performing rapid screening of small molecules.


Assuntos
Aflatoxina B1 , Anticorpos , Aflatoxina B1/análise , Antígenos , Arachis/metabolismo , Imunoensaio/métodos , Limite de Detecção
3.
Transpl Int ; 35: 10465, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35935272

RESUMO

For the past decades, complement activation and complement-mediated destruction of allograft cells were considered to play a central role in anti-HLA antibody-mediated rejection (AMR) of kidney transplants. However, also complement-independent mechanisms are relevant in the downstream immune activation induced by donor-specific antibodies, such as Fc-gamma receptor (FcγR)-mediated direct cellular activation. This article reviews the literature regarding FcγR involvement in AMR, and the potential contribution of FcγR gene polymorphisms to the risk for antibody mediated rejection of kidney transplants. There is large heterogeneity between the studies, both in the definition of the clinical phenotypes and in the technical aspects. The study populations were generally quite small, except for two larger study cohorts, which obviates drawing firm conclusions regarding the associations between AMR and specific FcγR polymorphisms. Although FcγR are central in the pathophysiology of AMR, it remains difficult to identify genetic risk factors for AMR in the recipient's genome, independent of clinical risk factors, independent of the donor-recipient genetic mismatch, and in the presence of powerful immunosuppressive agents. There is a need for larger, multi-center studies with standardised methods and endpoints to identify potentially relevant FcγR gene polymorphisms that represent an increased risk for AMR after kidney transplantation.


Assuntos
Transplante de Rim , Anticorpos , Rejeição de Enxerto , Imunossupressores , Transplante de Rim/efeitos adversos , Receptores de IgG
4.
Exp Clin Transplant ; 20(7): 695-697, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35924746

RESUMO

Chronic active antibody-mediated rejection is the leading cause of kidney transplant failure. Although various immunosuppressive agents have been tested, rituximab included, presently there is no effective treatment. There are reports about the beneficial role of certain immunosuppressive protocols that include rituximab to reduce donor-specific antibodies, the cause of chronic active antibody-mediated rejection. If an immunosuppressive agent reduces donor-specific antibodies, its administration before the occurrence of chronic active antibody-mediated rejection may be beneficial. We describe a case of a renaltransplantrecipient with recurrent membranous nephropathy and recent development of donor-specific antibodies but without histological evidence of active antibody-mediated rejection. The patient received 3 weekly doses of rituximab for recurrent membranous nephropathy, and complete remission was achieved. One year after, he has preserved an excellentrenal function without proteinuria. However, repeated measurements of donor-specific antibodies revealed that rituximab only modestly reduced donor-specific antibodies. Donor-specific antibody levels remained considerably higher than the laboratory reference value. Thus,rituximab alone may not have a role to prevent chronic active antibody- mediated rejection in patients with donor-specific antibodies.


Assuntos
Glomerulonefrite Membranosa , Imunossupressores , Transplante de Rim , Rituximab , Anticorpos , Glomerulonefrite Membranosa/diagnóstico , Glomerulonefrite Membranosa/tratamento farmacológico , Glomerulonefrite Membranosa/patologia , Rejeição de Enxerto/tratamento farmacológico , Rejeição de Enxerto/etiologia , Rejeição de Enxerto/prevenção & controle , Humanos , Imunossupressores/efeitos adversos , Transplante de Rim/efeitos adversos , Masculino , Rituximab/uso terapêutico , Resultado do Tratamento
5.
Sci Immunol ; 7(74): eade1493, 2022 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-35930656

RESUMO

Cytomegalovirus-infected moms with antibodies that can engage the immune system via FcγR are less likely to congenitally transmit the virus.


Assuntos
Infecções por Citomegalovirus , Citomegalovirus , Anticorpos , Humanos
6.
Transpl Int ; 35: 10626, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35928347

RESUMO

Alloimmune responses in kidney transplant (KT) patients previously hospitalized with COVID-19 are understudied. We analyzed a cohort of 112 kidney transplant recipients who were hospitalized following a positive SARS-CoV-2 test result during the first 20 months of the COVID-19 pandemic. We found a cumulative incidence of 17% for the development of new donor-specific antibodies (DSA) or increased levels of pre-existing DSA in hospitalized SARS-CoV-2-infected KT patients. This risk extended 8 months post-infection. These changes in DSA status were associated with late allograft dysfunction. Risk factors for new or increased DSA responses in this KT patient cohort included the presence of circulating DSA pre-COVID-19 diagnosis and time post-transplantation. COVID-19 vaccination prior to infection and remdesivir administration during infection were each associated with decreased likelihood of developing a new or increased DSA response. These data show that new or enhanced DSA responses frequently occur among KT patients requiring admission with COVID-19 and suggest that surveillance, vaccination, and antiviral therapies may be important tools to prevent alloimmunity in these individuals.


Assuntos
COVID-19 , Transplante de Rim , Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Anticorpos , COVID-19/tratamento farmacológico , COVID-19/prevenção & controle , Teste para COVID-19 , Vacinas contra COVID-19/uso terapêutico , Rejeição de Enxerto , Antígenos HLA , Humanos , Pandemias , SARS-CoV-2 , Transplantados , Vacinação
7.
J Vis Exp ; (185)2022 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-35938827

RESUMO

Arginine methylation is emerging as a key post-translational modification involved in a large range of biological processes. Its study in tissue is often limited by the lack of a specific antibody recognizing the target arginine residue. Proximity ligation assay (PLA) was originally developed to study protein/protein interactions. Here, we describe in detail a PLA protocol dedicated to the detection of arginine methylation that we applied to the glucocorticoid receptor (GR). Having previously shown that PRMT5 dimethylates GRs in cells, we used PLA with a pan symmetrical dimethyl antibody and an anti-GR antibody to measure GR methylation in breast tumors. We demonstrate that PLA offers a unique approach to measure arginine methylation of a target protein, even when the site of methylation has not been identified. This technique could be extended to other post-translational modifications where effective pan antibodies are available. Hence, we detail the PLA technology used to detect arginine methylation in fixed tissue using GR as an example.


Assuntos
Arginina , Fenômenos Biológicos , Anticorpos/metabolismo , Arginina/metabolismo , Metilação , Processamento de Proteína Pós-Traducional , Proteína-Arginina N-Metiltransferases/genética , Proteína-Arginina N-Metiltransferases/metabolismo , Proteínas/metabolismo
8.
Front Immunol ; 13: 939907, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35935998

RESUMO

Regulation of innate immune responses is essential for maintenance of immune homeostasis and development of an appropriate immunity against microbial infection. We show here that miR-3614-5p, product of the TRIM25 host gene, is induced by type I interferon (IFN-I) in several human non-immune and immune cell types, in particular in primary myeloid cells. Studies in HeLa cells showed that miR-3614-5p represses both p110 and p150 ADAR1 and reduces constitutive and IFN-induced A-to-I RNA editing. In line with this, activation of innate sensors and expression of IFN-ß and the pro-inflammatory IL-6 are promoted. MiR-3614-5p directly targets ADAR1 transcripts by binding to one specific site in the 3'UTR. Moreover, we could show that endogenous miR-3614-5p is associated with Ago2 and targets ADAR1 in IFN-stimulated cells. Overall, we propose that, by reducing ADAR1, IFN-I-induced miR-3614-5p contributes to lowering the activation threshold of innate sensors. Our findings provide new insights into the role of miR-3614-5p, placing it as a potential fine tuner of dsRNA metabolism, cell homeostasis and innate immunity.


Assuntos
Adenosina Desaminase/metabolismo , Imunidade Inata , Interferon Tipo I , MicroRNAs , Proteínas de Ligação a RNA/metabolismo , Fatores de Transcrição/genética , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Anticorpos , Antivirais , Células HeLa , Humanos , MicroRNAs/genética , Isoformas de Proteínas , RNA de Cadeia Dupla
9.
Mikrochim Acta ; 189(8): 271, 2022 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-35789294

RESUMO

A signal-on sandwich-like electrochemical immunosensor was built for determination of cytokeratin 19 fragments 21-1 (CYFRA 21-1) in non-small cell lung cancer (NSCLC) by confining electroactive dye (e.g., methylene blue, MB) as a probe for amplifying signals. Specifically, core-shell gold@rhodium dendritic nanocrystals (Au@Rh DNCs) behaved as a substrate for primary antibody and accelerate interfacial electron transfer. Besides, hollow carbon spheres (HCSs) were subsequently modified with polydopamine (PDA) and PtPd nanoparticles for sequential integration of the secondary antibody and confinement of MB as a label, termed as MB/PtPd/PDA/HCSs for clarity. The built sensors showed a broad linear range (100 fg mL-1 ~ 100 ng mL-1) for detection of CYFRA 21-1 with an ultra-low detection limit (31.72 fg mL-1, S/N = 3), coupled with satisfactory performance in human serum samples. This work can be explored for assays of other proteins and provides some constructive insights for early and accurate diagnosis of NSCLC.


Assuntos
Técnicas Biossensoriais , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Nanopartículas , Anticorpos , Antígenos de Neoplasias , Carbono , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Humanos , Imunoensaio , Indóis , Queratina-19 , Neoplasias Pulmonares/diagnóstico , Polímeros
11.
Lab Chip ; 22(16): 3055-3066, 2022 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-35851596

RESUMO

Personalized diagnostics of infectious diseases require monitoring disease progression due to their ever-changing physiological conditions and the multi-faceted organ system mechanisms involved in disease pathogenesis. In such instances, the recommended clinical strategies involve multiplexing data collection from critical biomarkers related to a patient's conditions along with longitudinal frequent patient monitoring. Numerous detection technologies exist both in research and commercial settings to monitor these conditions, however, they fail to provide biomarker multiplexing ability with design and data processing simplicity. For a recently conceived multiplexing biomarker modality, this work demonstrates the use of electrically sensitive microparticles targeting and identifying membrane receptors on leukocytes using a single detection source, with a high potential for multiplexing greater than any existing impedance-based single-detection scheme. Here, polystyrene microparticles are coated with varying thicknesses of metal oxides, which generate quantifiable impedance shifts when exposed to multifrequency electric fields depending on the metal oxide thickness. Using multifrequency impedance cytometry, these particles can be measured and differentiated rapidly across one coplanar electrode scheme. After surface-functionalizing particles with antibodies targeting CD11b and CD66b receptors, the particles are combined with isolated neutrophils to measure receptor expression. A combination of data analysis techniques including multivariate analysis, supervised machine learning, and unsupervised machine learning was able to accurately differentiate samples with up to 91% accuracy. This proof-of-concept study demonstrates the potential for these oxide-coated particles for enumerating specific leukocytes enabling multiplexing. Further, additional coating thicknesses or different metal oxide materials can enable a compendium of multiplexing targeting resource to be used to develop a high-multiplexing sensor for targeting membrane receptor expression.


Assuntos
Técnicas Analíticas Microfluídicas , Microfluídica , Óxido de Alumínio , Anticorpos , Biomarcadores , Impedância Elétrica , Humanos , Neutrófilos , Óxidos
12.
Histopathology ; 81(3): 389-401, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35791778

RESUMO

OBJECTIVE: Chondroblastoma-like osteosarcoma (CBLOS) is a rare and poorly understood variant of OS. We examined the clinicopathological, immunohistochemical and molecular features of six CBLOSs to highlight the differences with conventional high-grade OS (CHGOS) and CB, including CB with aggressive features. METHODS: We performed histone 3.3 mutation analysis by gene sequencing and/or immunohistochemistry in all cases, while whole exome sequencing (WES) was performed on two CB-like osteosarcomas and 11 conventional high-grade OS. RESULTS: CBLOSs were predominantly localised at acral sites and involved mainly male subjects with a mean age of 29 years. One patient who had metastases at presentation died of disease, while another patient who developed multiple local recurrences and lung metastases was alive with no evidence of disease (ANED) at 294 months. The remaining patients were ANED after a mean interval of 70.8 months. Histologically, all CBLOS presented aggressive features, including nuclear atypia and infiltrative growth. Immunohistochemistry with H3F3 K36M mutant antibody was negative in all CBLOSs, and none of the five tumours tested by gene sequencing had H3F3B mutations. Conversely, all CBs presented the H3F3B K36M variant and were positive for immunostaining with the H3F3 K36M antibody. Two CBLOSs analysed by WES differed in amount and type of mutation from 11 cases of CHGOS. Moreover, CBLOSs showed lower copy number alteration (CNA) score values than CHGOSs. CONCLUSIONS: CBLOS presents a different genetic background and a less aggressive clinical behaviour in comparison with CHGOS. Search of the H3F3B K36M mutation is useful in the differential diagnosis with CB.


Assuntos
Neoplasias Ósseas , Condroblastoma , Osteossarcoma , Adulto , Anticorpos , Neoplasias Ósseas/patologia , Condroblastoma/diagnóstico , Condroblastoma/genética , Condroblastoma/patologia , Histonas/genética , Humanos , Imuno-Histoquímica , Masculino , Osteossarcoma/patologia
13.
Ticks Tick Borne Dis ; 13(5): 101999, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35816827

RESUMO

Anaplasma phagocytophilum and Babesia microti are emerging tick-borne pathogens in the United States. Although active infection is typically diagnosed by direct diagnostic tests, such as blood smear or polymerase chain reaction assay, serologic assays can be helpful to identify past infections, and the use of acute plus convalescent testing can potentially identify recent infections. We employed a peptide array to select sets of linear peptides for serologic diagnosis of infections with A. phagocytophilum and B. microti. Three optimal peptides were selected for each agent based on their performance with clinical specimens. All three A. phagocytophilum peptides were located within the conserved fragments of the MSP2 antigen. Two B. microti peptides were located in the N terminus of the SA-1 antigen; the third was in the BMN 1-17 antigen. We found that these peptides can be a useful tool for detection of antibody reactivity to both of these pathogens.


Assuntos
Anaplasma phagocytophilum , Babesia microti , Babesiose , Borrelia burgdorferi , Anticorpos , Babesiose/diagnóstico , Humanos , Peptídeos
14.
Analyst ; 147(16): 3732-3740, 2022 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-35833583

RESUMO

Exosomes are vesicles released by healthy and cancer cells into the extracellular matrix and bodily fluid. Cancer cell-derived exosomes have attracted much attention in early-stage detection and prognostication of treatment response. Thus, detecting exosomes is of great interest to biology and medicine. However, many conventional detection methods require high-cost equipment and centralized laboratory facilities, making diagnostics inaccessible in limited-resource settings. This study reports a proof-of-concept low-cost electrochemical paper-based analytical device to quantify both the total bulk and cancer cell-derived exosomes in cell culture media. The device employs a sandwich immune assay design, where exosomes are initially captured using the electrode-bound generic antibodies (i.e. CD9) and subsequently detected via ovarian cancer-specific CA125 antibodies. Our proposed device quantifies the total bulk exosome concentration with a detection limit of 9.3 × 107 exosomes per mL and ovarian cancer cell-derived exosomes with a detection limit of 7.1 × 108 exosomes per mL, with a relative standard deviation of <10% (n = 3). We suggest that this low-cost and simple electrochemical paper-based device could be an alternative tool for detecting disease-specific exosomes in biological samples with the potential to be further developed for point-of-care diagnosis.


Assuntos
Exossomos , Neoplasias Ovarianas , Anticorpos , Eletrodos , Feminino , Humanos , Neoplasias Ovarianas/diagnóstico
15.
J Neuroimmunol ; 370: 577918, 2022 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-35853356

RESUMO

OBJECTIVES: To report two pediatric cases of autoimmune cerebellar ataxia associated with the anti-Leucine-rich glioma-inactivated protein 1 (LGI1)antibodies. METHODS: The clinical features of the two patients were described retrospectively. The indirect immunofluorescence using transfected cells (cell-based assay, CBA) and the rat cerebellum (tissue-based assay, TBA) with the multi-antigen co-plate biochip mosaic techniques were used to detect the antibodies. Clinical and laboratory characteristics were described. RESULTS: Two males were included. The onset ages were 2.7y and 4y, respectively. Patient 1 manifested as isolated acute cerebellar ataxia. Patient 2 had extra-cerebellar symptoms, including seizures, encephalopathy, faciobrachial dystonic seizures(FBDs), and significant cerebellar ataxia. The hyponatremia and tumors were not found. Both of them responded well to the immunotherapy. CONCLUSIONS: The autoimmune cerebellar ataxia might be a new phenotype of LGI1-Abs autoimmunity in children.


Assuntos
Ataxia Cerebelar , Glioma , Encefalite Límbica , Anticorpos , Autoanticorpos , Ataxia Cerebelar/complicações , Glioma/complicações , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Leucina/uso terapêutico , Masculino , Estudos Retrospectivos , Convulsões/tratamento farmacológico
16.
Anal Chem ; 94(31): 10967-10975, 2022 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-35895913

RESUMO

We present a method for monitoring spatially localized antigen-antibody binding events on physiologically relevant substrates (cell and tissue sections) using fluorescence lifetime imaging. Specifically, we use the difference between the fluorescence decay times of fluorescently tagged antibodies in free solution and in the bound state to track the bound fraction over time and hence deduce the binding kinetics. We make use of a microfluidic probe format to minimize the mass transport effects and localize the analysis to specific regions of interest on the biological substrates. This enables measurement of binding constants (kon) on surface-bound antigens and on cell blocks using model biomarkers. Finally, we directly measure p53 kinetics with differential biomarker expression in ovarian cancer tissue sections, observing that the degree of expression corresponds to the changes in kon, with values of 3.27-3.50 × 103 M-1 s-1 for high biomarker expression and 2.27-2.79 × 103 M-1 s-1 for low biomarker expression.


Assuntos
Neoplasias Ovarianas , Anticorpos , Reações Antígeno-Anticorpo , Feminino , Humanos , Cinética , Imagem Óptica
17.
Protein Eng Des Sel ; 352022 Feb 17.
Artigo em Inglês | MEDLINE | ID: mdl-35871543

RESUMO

TCR-like antibodies represent a unique type of engineered antibodies with specificity toward pHLA, a ligand normally restricted to the sensitive recognition by T cells. Here, we report a phage display-based sequential development path of such antibodies. The strategy goes from initial lead identification through in silico informed CDR engineering in combination with framework engineering for affinity and thermostability optimization, respectively. The strategy allowed the identification of HLA-DQ2.5 gluten peptide-specific TCR-like antibodies with low picomolar affinity. Our method outlines an efficient and general method for development of this promising class of antibodies, which should facilitate their utility including translation to human therapy.


Assuntos
Anticorpos , Bacteriófagos , Humanos , Peptídeos/genética , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T
18.
Mol Genet Metab ; 136(4): 249-259, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35835061

RESUMO

PURPOSE: Mucopolysaccharidosis IIIA (MPS IIIA) is an inherited lysosomal storage disorder caused by mutations in the N-sulfoglucosamine sulfohydrolase gene that result in deficient enzymatic degradation of heparan sulfate (HS), resulting in progressive neurodegeneration in early childhood and premature death. A chemically modified variant of recombinant human sulfamidase, SOBI003, has shown to cross the blood-brain barrier (BBB) in mice and achieve pharmacologically relevant levels in cerebrospinal fluid (CSF). We report on a phase 1/2, open-label, first-in-human (FIH) study (NCT03423186) and its extension study (NCT03811028) to evaluate the long-term safety, tolerability, pharmacokinetics/pharmacodynamics (PK/PD) and clinical efficacy of SOBI003 in patients with MPS IIIA for up to 104 weeks. METHODS: Six patients aged 1-6 years with confirmed MPS IIIA with developmental age ≥ 12 months received weekly intravenous injections of SOBI003 at 3 mg/kg (Cohort 1, n = 3) or 10 mg/kg (Cohort 2, n = 3). During the extension study, the individual dose of SOBI003 could be adjusted up to 20 mg/kg at the discretion of the investigator. RESULTS: SOBI003 was generally well tolerated. Serum concentrations of SOBI003 increased in proportion to dose, and presence in CSF confirmed that SOBI003 crosses the BBB. Anti-drug antibodies (ADA) were detected in serum and CSF in all patients, with subsequent reductions in serum SOBI003 exposure at high ADA titers. SOBI003 exerted a clear PD effect: a mean reduction in HS levels in CSF of 79% was recorded at the last assessment, together with reductions in HS levels in serum and urine. Neurocognitive development age-equivalent scores showed a stabilization of cognition for all patients, whereas no clear overall clinical effect was observed on adaptive behavior, sleep pattern or quality of life. CONCLUSION: SOBI003 was well tolerated when administered as weekly intravenous infusions at doses of up to 20 mg/kg for up to 104 weeks. ADA development was common and likely affected both PK and PD parameters. SOBI003 crossed the BBB and showed pharmacological activity on HS in CSF.


Assuntos
Mucopolissacaridose III , Anticorpos , Encéfalo/metabolismo , Criança , Pré-Escolar , Heparitina Sulfato/metabolismo , Humanos , Hidrolases , Lactente , Mucopolissacaridose III/tratamento farmacológico , Mucopolissacaridose III/genética , Qualidade de Vida
19.
J Neuroimmunol ; 370: 577932, 2022 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-35853357

RESUMO

A significant proportion of multiple sclerosis (MS) patients treated with interferon beta-1a (Rebif™) develop anti-drug antibodies (ADA) with a negative impact on treatment efficacy. We hypothesized that high-throughput B-cell receptor (BCR) repertoire analysis could be used to predict and monitor ADA development. To study this we analyzed 228 peripheral blood samples from 68 longitudinally followed patients starting on interferon beta-1a. Our results show that whole blood BCR analysis does not reflect, and does not predict ADA development in MS patients treated with interferon beta-1a. We propose that BCR analysis of phenotypically selected cell subsets or tissues might be more informative.


Assuntos
Esclerose Múltipla , Anticorpos/imunologia , Humanos , Interferon beta-1a/efeitos adversos , Interferon beta-1a/uso terapêutico , Esclerose Múltipla/tratamento farmacológico , Receptores de Antígenos de Linfócitos B/sangue , Receptores de Antígenos de Linfócitos B/imunologia
20.
Molecules ; 27(14)2022 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-35889369

RESUMO

This work aims at understanding the attachment mechanisms and stability of proteins on a chromatography medium to develop more efficient functionalization methodologies, which can be exploited in affinity chromatography. In particular, the study was focused on the understanding of the attachment mechanisms of bovine serum albumin (BSA), used as a ligand model, and protein G on novel amine-modified alumina monoliths as a stationary phase. Protein G was used to develop a column for antibody purification. The results showed that, at lower protein concentrations (i.e., 0.5 to 1.0 mg·mL-1), protein attachment follows a 1st-order kinetics compatible with the presence of covalent binding between the monolith and the protein. At higher protein concentrations (i.e., up to 10 mg·mL-1), the data preferably fit a 2nd-order kinetics. Such a change reflects a different mechanism in the protein attachment which, at higher concentrations, seems to be governed by physical adsorption resulting in a multilayered protein formation, due to the presence of ligand aggregates. The threshold condition for the prevalence of physical adsorption of BSA was found at a concentration higher than 1.0 mg·mL-1. Based on this result, protein concentrations of 0.7 and 1.0 mg·mL-1 were used for the functionalization of monoliths with protein G, allowing a maximum attachment of 1.43 mg of protein G/g of monolith. This column was then used for IgG binding-elution experiments, which resulted in an antibody attachment of 73.5% and, subsequently, elution of 86%, in acidic conditions. This proved the potential of the amine-functionalized monoliths for application in affinity chromatography.


Assuntos
Anticorpos , Soroalbumina Bovina , Adsorção , Aminas , Cromatografia de Afinidade/métodos , Ligantes
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