RESUMEN
BACKGROUND: Infection by SARS-CoV2 has become a challenge, especially for immunocompromised patients who show a weaker humoral response to COVID-19 vaccine. Tixagevimab+cilgavimab (Evusheld) is a combination of human monoclonal antibodies that can be used for pre-exposure prophylaxis to prevent infection or disease by SARS-CoV2. OBJECTIVES: Our study aimed to investigate the effectiveness of Evusheld by comparing an Exposed and an Unexposed group. STUDY DESIGN: Immunocompromised patients were enrolled in the Evusheld Group between March and September 2022. All patients had anti-spike IgG antibody levels <260 BAU/ml before administration of Evusheld. Blood samples for serological evaluations were collected, and anti-Spike antibodies were tested. For the Unexposed Group, a serologic test was performed at enrollment and a questionnaire was performed after 6 months. RESULTS: 43 patients received Evusheld pre-exposure prophylaxis and 45 patients not receiving Evusheld were enrolled in the Unexposed group. The median age was 59.0 years in the Evusheld group, and 63.0 in the unexposed group. In the Evusheld group, during the Omicron wave in Italy, 23.3% of subjects developed symptomatic infection compared to 42.2% in the unexposed group. A majority of infections was seen in male respect to female patients. No difference in length of infection between the groups was seen. Antibody level remained higher than the basal threshold at 180 days from enrollment. CONCLUSIONS: Evusheld seems to reduce the rate of symptomatic infection in immunocompromised patients. Further data are required to determine whether this prophylaxis may have a longer-lasting effect over time.
Asunto(s)
Anticuerpos Antivirales , Vacunas contra la COVID-19 , COVID-19 , Huésped Inmunocomprometido , Profilaxis Pre-Exposición , SARS-CoV-2 , Humanos , Masculino , Femenino , Persona de Mediana Edad , SARS-CoV-2/inmunología , COVID-19/prevención & control , COVID-19/inmunología , Vacunas contra la COVID-19/inmunología , Vacunas contra la COVID-19/administración & dosificación , Anciano , Profilaxis Pre-Exposición/métodos , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , Adulto , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Glicoproteína de la Espiga del Coronavirus/inmunología , Inmunoglobulina G/sangreRESUMEN
Aim: An assay to detect anti-tocilizumab antibodies in the presence of high levels of circulating target and drug is needed for immunogenicity assessment in comparative clinical studies.Methods: An assay was developed and validated using a combination of blocking agents and dilutions to overcome target interference challenges.Results: No false-positive signal was detected in serum samples spiked with 350-500 ng/ml of IL-6 receptor. As low as 50 ng/ml of positive control antibodies could be detected in the presence of either 500 ng/ml of IL-6 or 250 µg/ml of the drug product. Assay also demonstrated high sensitivity, selectivity and precision.Conclusion: A robust, easy to perform immunogenicity assay was developed and validated for detecting anti-tocilizumab antibodies.
[Box: see text].
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Anticuerpos Monoclonales Humanizados , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/sangre , Humanos , Anticuerpos/inmunología , Interleucina-6/sangre , Interleucina-6/inmunología , Inmunoensayo/métodosRESUMEN
The optimal efficacy of xenogeneically generated proteins intended for application in humans requires that their own antigenicity be minimized. This necessary adaptation of antibodies to a humanized version poses challenges since modifications even distant from the binding sites can greatly influence antigen recognition and this is the primary feature that must be maintained during all modifications. Current strategies often rely on grafting and/or randomization/selection to arrive at a humanized variant retaining the binding properties of the original molecule. However, in terms of speed and efficiency, rationally directed approaches can be superior, provided the requisite structural information is available. We present here a humanization procedure based on the high-resolution X-ray structure of a chimaeric IgG against a marker for multiple myeloma. Based on in silico modelling of humanizing amino acid substitutions identified from sequence alignments, we devised a straightforward cloning procedure to rapidly evaluate the proposed sequence changes. Careful inspection of the structure allowed the identification of a potentially problematic amino acid change that indeed disrupted antigen binding. Subsequent optimization of the antigen binding loop sequences resulted in substantial recovery of binding affinity lost in the completely humanized antibody. X-ray structures of the humanized and optimized variants demonstrate that the antigen binding mode is preserved, with surprisingly few direct contacts to antibody atoms. These results underline the importance of structural information for the efficient optimization of protein therapeutics. KEY MESSAGES: Structure-based humanization of an IgG against BCMA, a marker for Multiple Myeloma. Identification of problematic mutations and unexpected modification sites. Structures of the modified IgG-antigen complexes verified predictions. Provision of humanized high-affinity IgGs against BCMA for therapeutic applications.
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Mieloma Múltiple , Humanos , Mieloma Múltiple/terapia , Mieloma Múltiple/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/química , Inmunoglobulina G/inmunología , Inmunoglobulina G/química , Modelos Moleculares , Cristalografía por Rayos X , Secuencia de Aminoácidos , Conformación Proteica , Unión ProteicaRESUMEN
Monitoring belimumab concentrations in patients can be a valuable tool for assessing treatment response and for personalizing drug doses. Various assay formats may be used to measure concentrations of therapeutic monoclonal antibodies. A particularly useful format involves the use of anti-idiotype monoclonal antibodies, selected to be highly specific to the antibody of interest. Here, we describe the development of a specific, high-affinity anti-idiotype antibody to belimumab, and the application of this antibody in a homologous sandwich ELISA to measure belimumab concentrations.
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Anticuerpos Antiidiotipos , Anticuerpos Monoclonales Humanizados , Monitoreo de Drogas , Ensayo de Inmunoadsorción Enzimática , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/inmunología , Humanos , Ensayo de Inmunoadsorción Enzimática/métodos , Monitoreo de Drogas/métodos , Anticuerpos Antiidiotipos/inmunología , Animales , Inmunosupresores/sangreRESUMEN
Measurement of anti-drug antibodies (ADA) to assess the incidence of ADA in a clinical trial is a critical step in immunogenicity assessment during the development of a protein therapeutic. We developed novel graphical approaches to illustrate clinical trial ADA data for the PD-L1 inhibitor atezolizumab (Tecentriq) that included a systematic analysis of the impact of the timing of ADA sampling and ADA assay drug tolerance on reported ADA incidence. We found that approaches used across the industry for ADA incidence analysis provide a limited view of immunogenicity in oncology studies, where ADA detection may be confounded by both drug dosage and patient attrition. Moreover, these approaches can miss important temporal information about the immune response. Our results demonstrated that the methodology of ADA assessment for the atezolizumab program was specifically designed to capture most ADA responses to ensure accurate reporting of ADA incidence. We further showed that the use of sparse sampling and/or ADA test methods with insufficient drug tolerance may result in a significant underreporting of ADA incidence. We conclude that the comparison of ADA incidence between different drugs can be highly misleading and that a test method with appropriate sensitivity in the presence of the drug and a clinical sampling scheme that is aligned with ADA responses to a drug is required to accurately report ADA incidence.
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Anticuerpos Monoclonales Humanizados , Humanos , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos/inmunología , Tolerancia a Medicamentos/inmunología , Inhibidores de Puntos de Control Inmunológico/inmunologíaRESUMEN
Novel engineered IL-2 agonists strive to increase the therapeutic window of aldesleukin (human IL-2) by increasing selectivity toward effector over regulatory T cells and reducing dose-limiting toxicities. Here we describe ANV419, an IL-2/anti-IL2 antibody fusion protein designed for selective IL-2 receptor ßγ (IL-2 Rßγ) activation by sterically hindering IL-2 from binding to IL-2 Rα. The fusion protein has an IL-2 connected to the light chain complementarity-determining region (CDR) domain of a humanized antibody that binds to IL-2 at the same epitope as IL-2 Rα. Optimization of the selectivity and pharmacological properties led to the selection of ANV419. ANV419 preferentially expands CD8+ T cells and natural killer (NK) cells over Tregs and can be safely administered at doses that elicit strong pharmacodynamic effects and efficacy in mouse tumor models. Its anti-tumor efficacy was enhanced when combined with programmed cell death protein 1 (PD-1) or cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) checkpoint inhibitors. ANV419 also enhances the NK cell killing capacity and increases tumor growth inhibition when used alongside trastuzumab in a Her-2+ xenograft mouse model. In cynomolgus monkeys, the estimated half-life of ANV419 is 24 h, and doses that induced sustained expansion of effector cells were well tolerated without the severe toxicities typically observed with high-dose IL-2. These data support the clinical development of ANV419 in solid tumors and hematological malignancies as monotherapy and in combination with checkpoint inhibitors or agents that induce antibody-dependent cellular cytotoxicity. ANV419 is currently in Phase 1/2 clinical development and may provide cancer patients with a wider therapeutic window than aldesleukin.
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Linfocitos T CD8-positivos , Interleucina-2 , Células Asesinas Naturales , Proteínas Recombinantes de Fusión , Animales , Células Asesinas Naturales/inmunología , Humanos , Interleucina-2/inmunología , Linfocitos T CD8-positivos/inmunología , Ratones , Proteínas Recombinantes de Fusión/farmacología , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/genética , Inmunoterapia/métodos , Macaca fascicularis , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/farmacología , Neoplasias/inmunología , Neoplasias/terapia , Neoplasias/tratamiento farmacológico , Ensayos Antitumor por Modelo de Xenoinjerto , Línea Celular Tumoral , FemeninoRESUMEN
Active and passive immunization is used in high-risk patients to prevent severe courses of COVID-19, but the impact of prophylactic neutralizing antibodies on the immune reaction to the mRNA vaccines has remained enigmatic. Here we show that CD4 T and B cell responses to Spikevax booster immunization are suppressed by the therapeutic antibodies Casirivimab and Imdevimab. B cell and T cell responses were significantly induced in controls but not in antibody-treated patients. The data indicates that humoral immunity, i. e. high levels of antibodies, negatively impacts reactive immunity, resulting in blunted cellular responses upon boosting. This argues for temporal separation of vaccination efforts; with active vaccination preferably applied before prophylactic therapeutic antibody treatment.
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Anticuerpos Neutralizantes , Anticuerpos Antivirales , Linfocitos B , Vacunas contra la COVID-19 , COVID-19 , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus , Humanos , COVID-19/prevención & control , COVID-19/inmunología , Linfocitos B/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Anticuerpos Antivirales/inmunología , Anticuerpos Antivirales/sangre , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/sangre , Vacunas contra la COVID-19/inmunología , SARS-CoV-2/inmunología , Persona de Mediana Edad , Masculino , Femenino , Vacunación , Adulto , Anciano , Linfocitos T CD4-Positivos/inmunología , Linfocitos T/inmunología , Inmunización Secundaria , Inmunidad Humoral , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéuticoAsunto(s)
Anticuerpos Monoclonales Humanizados , Enfermedad de Crohn , Subunidad p40 de la Interleucina-12 , Subunidad p19 de la Interleucina-23 , Ustekinumab , Animales , Humanos , Ratones , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Enfermedad de Crohn/diagnóstico , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/inmunología , Subunidad p40 de la Interleucina-12/antagonistas & inhibidores , Subunidad p40 de la Interleucina-12/inmunología , Subunidad p19 de la Interleucina-23/antagonistas & inhibidores , Subunidad p19 de la Interleucina-23/inmunología , Ustekinumab/inmunología , Ustekinumab/uso terapéutico , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
Introduction: Humanization is typically adopted to reduce the immunogenicity of murine antibodies generated by hybridoma technology when used in humans. Methods: Two different strategies of antibody humanization are popularly employed, including "complementarity determining region (CDR) grafting" and "framework (FR) shuffling" to humanize a murine antibody against human programmed death-1 (PD-1), XM PD1. In CDR-grafting humanization, the CDRs of XM PD-1, were grafted into the human FR regions with high homology to the murine FR counterparts, and back mutations of key residues were performed to retain the antigen-binding affinities. While in FR-shuffling humanization, a combinatorial library of the six murine CDRs in-frame of XM PD-1 was constructed to a pool of human germline FRs for high-throughput screening for the most favorable variants. We evaluated many aspects which were important during antibody development of the molecules obtained by the two methods, including antibody purity, thermal stability, binding efficacy, predicted humanness, and immunogenicity, along with T cell epitope prediction for the humanized antibodies. Results: While the ideal molecule was not achieved through CDR grafting in this particular instance, FR-shuffling proved successful in identifying a suitable candidate. The study highlights FR-shuffling as an effective complementary approach that potentially increases the success rate of antibody humanization. It is particularly noted for its accessibility to those with a biological rather than a computational background. Discussion: The insights from this comparison are intended to assist other researchers in selecting appropriate humanization strategies for drug development, contributing to broader application and understanding in the field.
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Regiones Determinantes de Complementariedad , Receptor de Muerte Celular Programada 1 , Animales , Humanos , Ratones , Receptor de Muerte Celular Programada 1/inmunología , Regiones Determinantes de Complementariedad/inmunología , Regiones Determinantes de Complementariedad/genética , Anticuerpos Monoclonales Humanizados/inmunología , Epítopos de Linfocito T/inmunologíaRESUMEN
The treatment options for multiple myeloma (MM) have undergone significant transformation with the advent of immunotherapy. Novel therapies that focus on tumor antigens now drive advances in MM research. Bispecific antibodies (bsAbs) leverage revolutionary advances in bioengineering techniques and embody the second generation of antibody-based tumor therapy. Recent studies on bsAbs in relapsed/refractory MM cases have revealed remarkable efficacy and acceptable safety profiles. The approval of elranatamab and teclistamab represents the next step in the development of bsAbs for the treatment of MM. This review article addresses the antigen targeting, efficacy, safety, and strategies in the application of bsAbs against treatment-resistant MM, with a focus on clinical trials and real-world data.
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Anticuerpos Biespecíficos , Mieloma Múltiple , Mieloma Múltiple/inmunología , Mieloma Múltiple/terapia , Anticuerpos Biespecíficos/uso terapéutico , Anticuerpos Biespecíficos/inmunología , Humanos , Inmunoterapia/métodos , Antígenos de Neoplasias/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/inmunología , Antineoplásicos Inmunológicos/uso terapéutico , Antineoplásicos Inmunológicos/inmunologíaRESUMEN
INTRODUCTION: When a new disease occurs, one of the most affordable remedies is drugs containing specific antibodies to this infectious agent. The use of such drugs is aimed at reducing the amount of the pathogen in the macroorganism and the associated reduction in the severity of the symptoms of the disease or recovery. The purpose of this review is to analyze the experience of using immunoglobulins and monoclonal antibodies in the treatment of COVID-19 patients during the pandemic. RESULTS AND CONCLUSION: The two main groups of medical protective agents that block the penetration of the SARS-CoV-2 virus into permissive cells are drugs obtained from blood plasma of convalescents (immunoglobulin) and human monoclonal antibodies. The first group of drugs in the treatment of COVID-19 includes blood plasma of convalescents, which can be successfully used for emergency prevention. The main disadvantage of using blood plasma convalescents is the difficulty of standardization due to the different content of specific antibodies in donors. Another disadvantage is the undesirable side effects in recipients that occur after plasma administration. An alternative approach to COVID-19 therapy is the use of humanized and genetically engineered human monoclonal antibodies against certain epitopes of the SARS-CoV-2 virus. For example, monoclonal antibodies against receptor-binding domain of the S-protein, which prevents the virus from entering permissive cells and interrupts the development of infection. The advantages of these drugs are their safety, high specific activity, and the possibility of standardization. However, the complexity of their production and high cost make them inaccessible for mass use in practical medicine.
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Anticuerpos Monoclonales , COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/inmunología , SARS-CoV-2/efectos de los fármacos , COVID-19/inmunología , COVID-19/terapia , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Monoclonales/inmunología , Anticuerpos Antivirales/inmunología , Anticuerpos Antivirales/uso terapéutico , Inmunoglobulinas/uso terapéutico , Inmunoglobulinas/inmunología , Tratamiento Farmacológico de COVID-19 , Sueroterapia para COVID-19 , Inmunización Pasiva , Pandemias , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/inmunología , Antivirales/uso terapéuticoRESUMEN
Atezolizumab (ATZ) is a human monoclonal antibody, which has been granted multiple approvals from the US Food and Drug Administration (FDA) for the immunotherapy of different types of cancer. This study describes the prototype of a time-resolved fluoroimmunoassay (TRFIA) for the quantitation of ATZ in plasma. The assay involved the non-competitive binding of ATZ to its specific antigen [programmed death-ligand 1 (PD-L1) protein]. The immune complex formed on the inner surface of the assay plate wells was quantified by anti-human secondary antibody labeled with a chelate of europium-ethylenediaminetetraacetic acid. The enhanced fluorescence signal was generated by an enhanced fluorescence solution composed of thenoyltrifluoroacetone, trioctylphosphine oxide, and Triton X-100. The conditions of the TRFIA were refined, and its optimum procedures were established. The assay was validated in accordance with the immunoassay validation guidelines, and all the validation parameters were acceptable. The working range of the assay was 20-1000 pg mL-1, and its limit of quantitation was 20 pg mL-1. The assay was applied to the quantitation of ATZ in plasma samples with satisfactory accuracy and precision. The proposed TRFIA has significant benefits over the existing methodologies for the quantitation of ATZ in clinical settings.
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Anticuerpos Monoclonales Humanizados , Fluoroinmunoensayo , Fluoroinmunoensayo/métodos , Humanos , Anticuerpos Monoclonales Humanizados/sangre , Anticuerpos Monoclonales Humanizados/química , Anticuerpos Monoclonales Humanizados/inmunología , Fluorescencia , Factores de TiempoRESUMEN
BACKGROUND: Several PD-1 antibodies approved as anti-cancer therapies work by blocking the interaction of PD-1 with its ligand PD-L1, thus restoring anti-cancer T cell activities. These PD-1 antibodies lack inter-species cross-reactivity, necessitating surrogate antibodies for preclinical studies, which may limit the predictability and translatability of the studies. RESULTS: To overcome this limitation, we have developed an inter-species cross-reactive PD-1 antibody, GNUV201, by utilizing an enhanced diversity mouse platform (SHINE MOUSE™). GNUV201 equally binds to human PD-1 and mouse PD-1, equally inhibits the binding of human PD-1/PD-L1 and mouse PD-1/PD-L1, and effectively suppresses tumor growth in syngeneic mouse models. The epitope of GNUV201 mapped to the "FG loop" of hPD-1, distinct from those of Keytruda® ("C'D loop") and Opdivo® (N-term). Notably, the structural feature where the protruding epitope loop fits into GNUV201's binding pocket supports the enhanced binding affinity due to slower dissociation (8.7 times slower than Keytruda®). Furthermore, GNUV201 shows a stronger binding affinity at pH 6.0 (5.6 times strong than at pH 7.4), which mimics the hypoxic and acidic tumor microenvironment (TME). This phenomenon is not observed with marketed antibodies (Keytruda®, Opdivo®), implying that GNUV201 achieves more selective binding to and better occupancy on PD-1 in the TME. CONCLUSIONS: In summary, GNUV201 exhibited enhanced affinity for PD-1 with slow dissociation and preferential binding in TME-mimicking low pH. Human/monkey/mouse inter-species cross-reactivity of GNUV201 could enable more predictable and translatable efficacy and toxicity preclinical studies. These results suggest that GNUV201 could be an ideal antibody candidate for anti-cancer drug development.
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Reacciones Cruzadas , Inmunoterapia , Receptor de Muerte Celular Programada 1 , Animales , Humanos , Receptor de Muerte Celular Programada 1/inmunología , Receptor de Muerte Celular Programada 1/metabolismo , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Ratones , Reacciones Cruzadas/inmunología , Inmunoterapia/métodos , Concentración de Iones de Hidrógeno , Neoplasias/inmunología , Neoplasias/terapia , Antígeno B7-H1/inmunología , Antígeno B7-H1/metabolismo , Antígeno B7-H1/antagonistas & inhibidores , Línea Celular Tumoral , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Inhibidores de Puntos de Control Inmunológico/farmacología , Epítopos/inmunología , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/farmacología , Ratones Endogámicos C57BL , FemeninoRESUMEN
One of the most important attributes of anti-amyloid antibodies is their selective binding to oligomeric and amyloid aggregates. However, current methods of examining the binding specificities of anti-amyloid ß (Aß) antibodies have limited ability to differentiate between complexes that form between antibodies and monomeric or oligomeric Aß species during the dynamic Aß aggregation process. Here, we present a high-resolution native ion-mobility mass spectrometry (nIM-MS) method to investigate complexes formed between a variety of Aß oligomers and three Aß-specific IgGs, namely two antibodies with relatively high conformational specificity (aducanumab and A34) and one antibody with low conformational specificity (crenezumab). We found that crenezumab primarily binds Aß monomers, while aducanumab preferentially binds Aß monomers and dimers and A34 preferentially binds Aß dimers, trimers, and tetrameters. Through collision induced unfolding (CIU) analysis, our data indicate that antibody stability is increased upon Aß binding and, surprisingly, this stabilization involves the Fc region. Together, we conclude that nIM-MS and CIU enable the identification of Aß antibody binding stoichiometries and provide important details regarding antibody binding mechanisms.
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Péptidos beta-Amiloides , Anticuerpos Monoclonales Humanizados , Espectrometría de Movilidad Iónica , Péptidos beta-Amiloides/química , Péptidos beta-Amiloides/inmunología , Péptidos beta-Amiloides/metabolismo , Anticuerpos Monoclonales Humanizados/química , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/metabolismo , Espectrometría de Movilidad Iónica/métodos , Humanos , Espectrometría de Masas/métodos , Unión Proteica , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Multimerización de ProteínaRESUMEN
Epitope binning, an approach for grouping antibodies based on epitope similarities, is a critical step in antibody drug discovery. However, conventional methods are complex, involving individual antibody production. Here, we established Epitope Binning-seq, an epitope binning platform for simultaneously analyzing multiple antibodies. In this system, epitope similarity between the query antibodies (qAbs) displayed on antigen-expressing cells and a fluorescently labeled reference antibody (rAb) targeting a desired epitope is analyzed by flow cytometry. The qAbs with epitope similar to the rAb can be identified by next-generation sequencing analysis of fluorescence-negative cells. Sensitivity and reliability of this system are confirmed using rAbs, pertuzumab and trastuzumab, which target human epidermal growth factor receptor 2. Epitope Binning-seq enables simultaneous epitope evaluation of 14 qAbs at various abundances in libraries, grouping them into respective epitope bins. This versatile platform is applicable to diverse antibodies and antigens, potentially expediting the identification of clinically useful antibodies.
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Epítopos , Humanos , Epítopos/inmunología , Análisis de Secuencia de ADN/métodos , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Animales , Receptor ErbB-2/inmunología , Receptor ErbB-2/genética , Citometría de Flujo/métodos , Trastuzumab/inmunología , Mapeo Epitopo/métodos , Anticuerpos/inmunología , Anticuerpos/genética , Anticuerpos Monoclonales Humanizados/inmunologíaRESUMEN
New therapies to treat or prevent viral infections are essential, as recently observed during the COVID-19 pandemic. Here, we propose a therapeutic strategy based on monoclonal antibodies that block the specific interaction between the host receptor Siglec-1/CD169 and gangliosides embedded in the viral envelope. Antibodies are an excellent option for treating infectious diseases based on their high specificity, strong targeting affinity, and relatively low toxicity. Through a process of humanization, we optimized monoclonal antibodies to eliminate sequence liabilities and performed biophysical characterization. We demonstrated that they maintain their ability to block viral entry into myeloid cells. These molecular improvements during the discovery stage are key if we are to maximize efforts to develop new therapeutic strategies. Humanized monoclonal antibodies targeting CD169 provide new opportunities in the treatment of infections caused by ganglioside-containing enveloped viruses, which pose a constant threat to human health. In contrast with current neutralizing antibodies that bind antigens on the infectious particle, our antibodies can prevent several types of enveloped viruses interacting with host cells because they target the host CD169 protein, thus becoming a potential pan-antiviral therapy.
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Anticuerpos Monoclonales Humanizados , Antivirales , Lectina 1 Similar a Ig de Unión al Ácido Siálico , Lectina 1 Similar a Ig de Unión al Ácido Siálico/inmunología , Humanos , Anticuerpos Monoclonales Humanizados/farmacología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/inmunología , Antivirales/farmacología , Antivirales/uso terapéutico , Animales , Tratamiento Farmacológico de COVID-19 , Internalización del Virus/efectos de los fármacos , SARS-CoV-2/inmunología , SARS-CoV-2/efectos de los fármacosRESUMEN
Two SARS-CoV-2 XBB sub-variants, FL.1 and GE.1, have been increasing in prevalence worldwide, but limited information is available about their ability to evade the immune system. FL.1 and GE.1 are emerging Omicron XBB variants possessing additional mutations in the spike RBD raising concerns of increased neutralization escape. In this study, we assessed the neutralizing ability of eleven FDA-approved monoclonal antibody combinations against different Omicron variants, including BA.2.75, BA.2.76, BA.4/5, XBB.1.5, and CH.1.1. Among the eleven antibodies, Sotrovimab was the only antibody to show broad neutralization ability against XBB.1.5. However, Sotrovimab showed attenuated neutralization efficiency against recently emerging XBB sub-lineages EG.5, FL.1, and GE.1 compared to XBB.1.5. Additionally, XBB.1.5 seropositive convalescent sera displayed lower neutralization activity against EG.5, FL.1, and GE.1. Overall, our findings present enhanced immune evasion capacity of emerging XBB variants and emphasize the importance of continued monitoring of novel variants.
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Anticuerpos Monoclonales , Anticuerpos Neutralizantes , Anticuerpos Antivirales , COVID-19 , Pruebas de Neutralización , SARS-CoV-2 , Humanos , SARS-CoV-2/inmunología , SARS-CoV-2/genética , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , COVID-19/inmunología , COVID-19/virología , Anticuerpos Monoclonales/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/genética , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/uso terapéutico , MutaciónRESUMEN
CD40 is a member of the tumor necrosis factor receptor superfamily, and it is widely expressed on immune and non-immune cell types. The interaction between CD40 and the CD40 ligand (CD40L) plays an essential function in signaling, and the CD40/CD40L complex works as an immune checkpoint molecule. CD40 has become a therapeutic target, and a variety of agonistic/antagonistic anti-CD40 monoclonal antibodies (mAbs) have been developed. To better understand the mode of action of anti-CD40 mAbs, we determined the X-ray crystal structures of dacetuzumab (agonist) and bleselumab (antagonist) in complex with the extracellular domain of human CD40, respectively. The structure reveals that dacetuzumab binds to CD40 on the top of cysteine-rich domain 1 (CRD1), which is the domain most distant from the cell surface, and it does not compete with CD40L binding. The binding interface of bleselumab spread between CRD2 and CRD1, overlapping with the binding surface of the ligand. Our results offer important insights for future structural and functional studies of CD40 and provide clues to understanding the mechanism of biological response. These data can be applied to developing new strategies for designing antibodies with more therapeutic efficacy.
Asunto(s)
Anticuerpos Monoclonales Humanizados , Antígenos CD40 , Humanos , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales Humanizados/química , Anticuerpos Monoclonales Humanizados/inmunología , Sitios de Unión , Antígenos CD40/química , Antígenos CD40/inmunología , Antígenos CD40/metabolismo , Ligando de CD40/química , Ligando de CD40/metabolismo , Ligando de CD40/inmunología , Cristalografía por Rayos X , Modelos Moleculares , Unión Proteica , Conformación ProteicaRESUMEN
The assembly of tissue-damaging membrane attack complexes (MACs; C5b-9) is a major mechanism by which excessive complement activation causes diseases. We previously developed a mouse anti-human C6 monoclonal antibody (mAb) 1C9 that selectively inhibits the assembly of MACs in human and non-human primates. In this project, we found that 1C9 also cross-reacted with rat and guinea pig C6, and determined its binding domains on C6 using different truncated C6 proteins. We then humanized the anti-C6 mAb by molecular modeling and complementarity-determining region grafting. After screening a library of 276 humanized variants with different combinations of humanized light and heavy chains in biophysical assays, we identified clone 3713 with the best developability profile, and an increased affinity against C6 when compared with the parental 1C9 mAb. This humanized 3713 mAb inhibited human, monkey, and rat complement-mediated hemolysis in vitro, and more importantly, it significantly reduced complement-mediated hemolysis in vivo in rats. These results demonstrated the successful humanization of the anti-C6 mAb and suggested that the humanized 3713 mAb could be further developed as a new therapeutic that selectively targets MAC for certain complement-mediated pathological conditions.
Asunto(s)
Anticuerpos Monoclonales , Complemento C6 , Hemólisis , Animales , Humanos , Ratas , Cobayas , Ratones , Hemólisis/efectos de los fármacos , Hemólisis/inmunología , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Complemento C6/inmunología , Anticuerpos Monoclonales Humanizados/inmunología , Anticuerpos Monoclonales Humanizados/farmacología , Activación de Complemento/inmunología , Activación de Complemento/efectos de los fármacos , Complejo de Ataque a Membrana del Sistema Complemento/inmunología , Reacciones Cruzadas/inmunologíaRESUMEN
TIGIT is mainly expressed on T cells and is an inhibitory checkpoint receptor that binds to its ligand PVR in the tumor microenvironment. Anti-TIGIT monoclonal antibodies (mAbs) such as Ociperlimab and Tiragolumab block the TIGIT-PVR interaction and are in clinical development. However, the molecular blockade mechanism of these mAbs remains elusive. Here, we report the crystal structures of TIGIT in complex with Ociperlimab_Fab and Tiragolumab_Fab revealing that both mAbs bind TIGIT with a large steric clash with PVR. Furthermore, several critical epitopic residues are identified. Interestingly, the binding affinity of Ociperlimab toward TIGIT increases approximately 17-fold when lowering the pH from 7.4 to 6.0. Our structure shows a strong electrostatic interaction between ASP103HCDR3 and HIS76TIGIT explaining the pH-responsive mechanism of Ociperlimab. In contrast, Tiragolumab does not show an acidic pH-dependent binding enhancement. Our results provide valuable information that could help to improve the efficacy of therapeutic antibodies for cancer treatment.