RESUMO
Most human monoclonal antibodies (mAbs) neutralizing SARS-CoV-2 recognize the spike (S) protein receptor-binding domain and block virus interactions with the cellular receptor angiotensin-converting enzyme 2. We describe a panel of human mAbs binding to diverse epitopes on the N-terminal domain (NTD) of S protein from SARS-CoV-2 convalescent donors and found a minority of these possessed neutralizing activity. Two mAbs (COV2-2676 and COV2-2489) inhibited infection of authentic SARS-CoV-2 and recombinant VSV/SARS-CoV-2 viruses. We mapped their binding epitopes by alanine-scanning mutagenesis and selection of functional SARS-CoV-2 S neutralization escape variants. Mechanistic studies showed that these antibodies neutralize in part by inhibiting a post-attachment step in the infection cycle. COV2-2676 and COV2-2489 offered protection either as prophylaxis or therapy, and Fc effector functions were required for optimal protection. Thus, natural infection induces a subset of potent NTD-specific mAbs that leverage neutralizing and Fc-mediated activities to protect against SARS-CoV-2 infection using multiple functional attributes.
Assuntos
Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/farmacologia , Substâncias Protetoras/farmacologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Ligação Competitiva , COVID-19/imunologia , COVID-19/virologia , Quimiocinas/metabolismo , Chlorocebus aethiops , Células HEK293 , Humanos , Fragmentos Fab das Imunoglobulinas/metabolismo , Imunoglobulina G/metabolismo , Pulmão/metabolismo , Camundongos Endogâmicos C57BL , Modelos Moleculares , Mutagênese/genética , Testes de Neutralização , Domínios Proteicos , Células VeroRESUMO
A safe and controlled manipulation of endocytosis in vivo may have disruptive therapeutic potential. Here, we demonstrate that the anti-emetic/anti-psychotic prochlorperazine can be repurposed to reversibly inhibit the in vivo endocytosis of membrane proteins targeted by therapeutic monoclonal antibodies, as directly demonstrated by our human tumor ex vivo assay. Temporary endocytosis inhibition results in enhanced target availability and improved efficiency of natural killer cell-mediated antibody-dependent cellular cytotoxicity (ADCC), a mediator of clinical responses induced by IgG1 antibodies, demonstrated here for cetuximab, trastuzumab, and avelumab. Extensive analysis of downstream signaling pathways ruled out on-target toxicities. By overcoming the heterogeneity of drug target availability that frequently characterizes poorly responsive or resistant tumors, clinical application of reversible endocytosis inhibition may considerably improve the clinical benefit of ADCC-mediating therapeutic antibodies.
Assuntos
Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/imunologia , Neoplasias/tratamento farmacológico , Proclorperazina/farmacologia , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais Humanizados/farmacologia , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Apresentação de Antígeno/efeitos dos fármacos , Biópsia , Cetuximab/farmacologia , Sistemas de Liberação de Medicamentos/métodos , Resistencia a Medicamentos Antineoplásicos/genética , Endocitose/efeitos dos fármacos , Endocitose/imunologia , Xenoenxertos , Humanos , Imunoglobulina G/genética , Imunoglobulina G/imunologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Células MCF-7 , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Neoplasias/genética , Neoplasias/imunologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Trastuzumab/farmacologiaRESUMO
Interleukin (IL)-1R3 is the co-receptor in three signaling pathways that involve six cytokines of the IL-1 family (IL-1α, IL-1ß, IL-33, IL-36α, IL-36ß and IL-36γ). In many diseases, multiple cytokines contribute to disease pathogenesis. For example, in asthma, both IL-33 and IL-1 are of major importance, as are IL-36 and IL-1 in psoriasis. We developed a blocking monoclonal antibody (mAb) to human IL-1R3 (MAB-hR3) and demonstrate here that this antibody specifically inhibits signaling via IL-1, IL-33 and IL-36 in vitro. Also, in three distinct in vivo models of disease (crystal-induced peritonitis, allergic airway inflammation and psoriasis), we found that targeting IL-1R3 with a single mAb to mouse IL-1R3 (MAB-mR3) significantly attenuated heterogeneous cytokine-driven inflammation and disease severity. We conclude that in diseases driven by multiple cytokines, a single antagonistic agent such as a mAb to IL-1R3 is a therapeutic option with considerable translational benefit.
Assuntos
Anticorpos Bloqueadores/farmacologia , Anticorpos Monoclonais/farmacologia , Proteína Acessória do Receptor de Interleucina-1/antagonistas & inibidores , Peritonite/imunologia , Pneumonia/imunologia , Psoríase/imunologia , Células A549 , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Células HEK293 , Humanos , Imiquimode/toxicidade , Inflamação/patologia , Interleucina-1/imunologia , Proteína Acessória do Receptor de Interleucina-1/imunologia , Interleucina-1beta/imunologia , Interleucina-33/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/toxicidade , Peritonite/tratamento farmacológico , Peritonite/patologia , Pneumonia/tratamento farmacológico , Pneumonia/patologia , Psoríase/tratamento farmacológico , Psoríase/patologia , Transdução de Sinais/imunologia , Ácido Úrico/toxicidadeRESUMO
Antibody responses during infection and vaccination typically undergo affinity maturation to achieve high-affinity binding for efficient neutralization of pathogens1,2. Similarly, high affinity is routinely the goal for therapeutic antibody generation. However, in contrast to naturally occurring or direct-targeting therapeutic antibodies, immunomodulatory antibodies, which are designed to modulate receptor signalling, have not been widely examined for their affinity-function relationship. Here we examine three separate immunologically important receptors spanning two receptor superfamilies: CD40, 4-1BB and PD-1. We show that low rather than high affinity delivers greater activity through increased clustering. This approach delivered higher immune cell activation, in vivo T cell expansion and antitumour activity in the case of CD40. Moreover, an inert anti-4-1BB monoclonal antibody was transformed into an agonist. Low-affinity variants of the clinically important antagonistic anti-PD-1 monoclonal antibody nivolumab also mediated more potent signalling and affected T cell activation. These findings reveal a new paradigm for augmenting agonism across diverse receptor families and shed light on the mechanism of antibody-mediated receptor signalling. Such affinity engineering offers a rational, efficient and highly tuneable solution to deliver antibody-mediated receptor activity across a range of potencies suitable for translation to the treatment of human disease.
Assuntos
Anticorpos Monoclonais , Afinidade de Anticorpos , Imunomodulação , Humanos , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Antígenos CD40/efeitos dos fármacos , Antígenos CD40/imunologia , Imunomodulação/efeitos dos fármacos , Imunomodulação/imunologia , Nivolumabe/imunologia , Nivolumabe/farmacologiaRESUMO
Epithelial-to-mesenchymal transition (EMT) regulates tumour initiation, progression, metastasis and resistance to anti-cancer therapy1-7. Although great progress has been made in understanding the role of EMT and its regulatory mechanisms in cancer, no therapeutic strategy to pharmacologically target EMT has been identified. Here we found that netrin-1 is upregulated in a primary mouse model of skin squamous cell carcinoma (SCC) exhibiting spontaneous EMT. Pharmacological inhibition of netrin-1 by administration of NP137, a netrin-1-blocking monoclonal antibody currently used in clinical trials in human cancer (ClinicalTrials.gov identifier NCT02977195 ), decreased the proportion of EMT tumour cells in skin SCC, decreased the number of metastases and increased the sensitivity of tumour cells to chemotherapy. Single-cell RNA sequencing revealed the presence of different EMT states, including epithelial, early and late hybrid EMT, and full EMT states, in control SCC. By contrast, administration of NP137 prevented the progression of cancer cells towards a late EMT state and sustained tumour epithelial states. Short hairpin RNA knockdown of netrin-1 and its receptor UNC5B in EPCAM+ tumour cells inhibited EMT in vitro in the absence of stromal cells and regulated a common gene signature that promotes tumour epithelial state and restricts EMT. To assess the relevance of these findings to human cancers, we treated mice transplanted with the A549 human cancer cell line-which undergoes EMT following TGFß1 administration8,9-with NP137. Netrin-1 inhibition decreased EMT in these transplanted A549 cells. Together, our results identify a pharmacological strategy for targeting EMT in cancer, opening up novel therapeutic interventions for anti-cancer therapy.
Assuntos
Anticorpos Monoclonais , Carcinoma de Células Escamosas , Transição Epitelial-Mesenquimal , Netrina-1 , Neoplasias Cutâneas , Animais , Humanos , Camundongos , Células A549 , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Receptores de Netrina/antagonistas & inibidores , Receptores de Netrina/deficiência , Receptores de Netrina/genética , Netrina-1/antagonistas & inibidores , Netrina-1/deficiência , Netrina-1/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Modelos Animais de Doenças , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Metástase Neoplásica/tratamento farmacológico , Análise da Expressão Gênica de Célula Única , RNA-Seq , Molécula de Adesão da Célula Epitelial/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Fator de Crescimento Transformador beta1/farmacologiaRESUMO
Resistance to checkpoint-blockade treatments is a challenge in the clinic. We found that although treatment with combined anti-CTLA-4 and anti-PD-1 improved control of established tumors, this combination compromised anti-tumor immunity in the low tumor burden (LTB) state in pre-clinical models as well as in melanoma patients. Activated tumor-specific T cells expressed higher amounts of interferon-γ (IFN-γ) receptor and were more susceptible to apoptosis than naive T cells. Combination treatment induced deletion of tumor-specific T cells and altered the T cell repertoire landscape, skewing the distribution of T cells toward lower-frequency clonotypes. Additionally, combination therapy induced higher IFN-γ production in the LTB state than in the high tumor burden (HTB) state on a per-cell basis, reflecting a less exhausted immune status in the LTB state. Thus, elevated IFN-γ secretion in the LTB state contributes to the development of an immune-intrinsic mechanism of resistance to combination checkpoint blockade, highlighting the importance of achieving the optimal magnitude of immune stimulation for successful combination immunotherapy strategies.
Assuntos
Anticorpos Monoclonais/farmacologia , Antígeno CTLA-4/antagonistas & inibidores , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Interferon gama/farmacologia , Neoplasias Experimentais/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Linfócitos T/efeitos dos fármacos , Animais , Anticorpos Monoclonais/imunologia , Antígeno CTLA-4/imunologia , Antígeno CTLA-4/metabolismo , Linhagem Celular Tumoral , Deleção Clonal/efeitos dos fármacos , Deleção Clonal/imunologia , Resistencia a Medicamentos Antineoplásicos/imunologia , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/metabolismo , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/imunologiaRESUMO
The recent emergence of SARS-CoV-2 Omicron (B.1.1.529 lineage) variants possessing numerous mutations has raised concerns of decreased effectiveness of current vaccines, therapeutic monoclonal antibodies and antiviral drugs for COVID-19 against these variants1,2. The original Omicron lineage, BA.1, prevailed in many countries, but more recently, BA.2 has become dominant in at least 68 countries3. Here we evaluated the replicative ability and pathogenicity of authentic infectious BA.2 isolates in immunocompetent and human ACE2-expressing mice and hamsters. In contrast to recent data with chimeric, recombinant SARS-CoV-2 strains expressing the spike proteins of BA.1 and BA.2 on an ancestral WK-521 backbone4, we observed similar infectivity and pathogenicity in mice and hamsters for BA.2 and BA.1, and less pathogenicity compared with early SARS-CoV-2 strains. We also observed a marked and significant reduction in the neutralizing activity of plasma from individuals who had recovered from COVID-19 and vaccine recipients against BA.2 compared to ancestral and Delta variant strains. In addition, we found that some therapeutic monoclonal antibodies (REGN10987 plus REGN10933, COV2-2196 plus COV2-2130, and S309) and antiviral drugs (molnupiravir, nirmatrelvir and S-217622) can restrict viral infection in the respiratory organs of BA.2-infected hamsters. These findings suggest that the replication and pathogenicity of BA.2 is similar to that of BA.1 in rodents and that several therapeutic monoclonal antibodies and antiviral compounds are effective against Omicron BA.2 variants.
Assuntos
Antivirais , Tratamento Farmacológico da COVID-19 , SARS-CoV-2 , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais Humanizados , Anticorpos Neutralizantes/farmacologia , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/farmacologia , Anticorpos Antivirais/uso terapêutico , Antivirais/farmacologia , Antivirais/uso terapêutico , COVID-19/genética , COVID-19/imunologia , COVID-19/virologia , Cricetinae , Citidina/análogos & derivados , Combinação de Medicamentos , Hidroxilaminas , Indazóis , Lactamas , Leucina , Camundongos , Nitrilas , Prolina , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/genética , SARS-CoV-2/patogenicidade , Glicoproteína da Espícula de Coronavírus/genética , Triazinas , TriazóisRESUMO
Despite its outstanding clinical success, immune checkpoint blockade remains ineffective in many patients. Accordingly, combination therapy capable of achieving greater antitumor immunity is urgently required. Here, we report that limiting glutamine metabolism in cancer cells bolsters the effectiveness of anti-programmed death ligand-1 (PD-L1) antibody. Inhibition of glutamine utilization increased PD-L1 levels in cancer cells, thereby inactivating co-cultured T cells. Under glutamine-limited conditions, reduced cellular GSH levels caused an upregulation of PD-L1 expression by impairing SERCA activity, which activates the calcium/NF-κB signaling cascade. Consequently, in tumors grown in immunocompetent mice, inhibition of glutamine metabolism decreased the antitumor activity of T cells. In combination with anti-PD-L1, however, glutamine depletion strongly promoted the antitumor efficacy of T cells in vitro and in vivo due to simultaneous increases in Fas/CD95 levels. Our results demonstrate the relevance of cancer glutamine metabolism to antitumor immunity and suggest that co-targeting of glutamine metabolism and PD-L1 represents a promising therapeutic approach.
Assuntos
Anticorpos Monoclonais/farmacologia , Antígeno B7-H1/metabolismo , Glutamina/metabolismo , Glutationa/metabolismo , Neoplasias/imunologia , Neoplasias/prevenção & controle , Linfócitos T/imunologia , Idoso , Animais , Apoptose , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Proliferação de Células , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Nus , Neoplasias/metabolismo , Neoplasias/patologia , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/antagonistas & inibidores , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Ebolaviruses cause severe disease in humans, and identification of monoclonal antibodies (mAbs) that are effective against multiple ebolaviruses are important for therapeutics development. Here we describe a distinct class of broadly neutralizing human mAbs with protective capacity against three ebolaviruses infectious for humans: Ebola (EBOV), Sudan (SUDV), and Bundibugyo (BDBV) viruses. We isolated mAbs from human survivors of ebolavirus disease and identified a potent mAb, EBOV-520, which bound to an epitope in the glycoprotein (GP) base region. EBOV-520 efficiently neutralized EBOV, BDBV, and SUDV and also showed protective capacity in relevant animal models of these infections. EBOV-520 mediated protection principally by direct virus neutralization and exhibited multifunctional properties. This study identified a potent naturally occurring mAb and defined key features of the human antibody response that may contribute to broad protection. This multifunctional mAb and related clones are promising candidates for development as broadly protective pan-ebolavirus therapeutic molecules.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/farmacologia , Ebolavirus/imunologia , Glicoproteínas/imunologia , Doença pelo Vírus Ebola/imunologia , Células 3T3 , Adulto , Animais , Células CHO , Linhagem Celular , Chlorocebus aethiops , Cricetulus , Modelos Animais de Doenças , Drosophila , Feminino , Furões , Cobaias , Doença pelo Vírus Ebola/prevenção & controle , Doença pelo Vírus Ebola/virologia , Humanos , Imunoglobulina G/imunologia , Células Jurkat , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Células THP-1 , Células VeroRESUMO
The costimulatory receptor 4-1BB is expressed on activated immune cells, including activated T cells. Antibodies targeting 4-1BB enhance the proliferation and survival of antigen-stimulated T cells in vitro and promote CD8 T cell-dependent anti-tumor immunity in pre-clinical cancer models. We found that T regulatory (Treg) cells infiltrating human or murine tumors expressed high amounts of 4-1BB. Intra-tumoral Treg cells were preferentially depleted by anti-4-1BB mAbs in vivo. Anti-4-1BB mAbs also promoted effector T cell agonism to promote tumor rejection. These distinct mechanisms were competitive and dependent on antibody isotype and FcγR availability. Administration of anti-4-1BB IgG2a, which preferentially depletes Treg cells, followed by either agonistic anti-4-1BB IgG1 or anti-PD-1 mAb augmented anti-tumor responses in multiple solid tumor models. An antibody engineered to optimize both FcγR-dependent Treg cell depleting capacity and FcγR-independent agonism delivered enhanced anti-tumor therapy. These insights into the effector mechanisms of anti-4-1BB mAbs lay the groundwork for translation into the clinic.
Assuntos
Anticorpos Monoclonais/farmacologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Imunomodulação/efeitos dos fármacos , Neoplasias/imunologia , Neoplasias/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/antagonistas & inibidores , Animais , Expressão Gênica , Humanos , Imunoglobulina G/farmacologia , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Camundongos , Camundongos Knockout , Neoplasias/genética , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/genética , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismoRESUMO
Monoclonal antibodies with neutralizing activity against SARS-CoV-2 have demonstrated clinical benefits in cases of mild-to-moderate SARS-CoV-2 infection, substantially reducing the risk for hospitalization and severe disease1-4. Treatment generally requires the administration of high doses of these monoclonal antibodies and has limited efficacy in preventing disease complications or mortality among hospitalized patients with COVID-195. Here we report the development and evaluation of anti-SARS-CoV-2 monoclonal antibodies with optimized Fc domains that show superior potency for prevention or treatment of COVID-19. Using several animal disease models of COVID-196,7, we demonstrate that selective engagement of activating Fcγ receptors results in improved efficacy in both preventing and treating disease-induced weight loss and mortality, significantly reducing the dose required to confer full protection against SARS-CoV-2 challenge and for treatment of pre-infected animals. Our results highlight the importance of Fcγ receptor pathways in driving antibody-mediated antiviral immunity and exclude the possibility of pathogenic or disease-enhancing effects of Fcγ receptor engagement of anti-SARS-CoV-2 antibodies upon infection. These findings have important implications for the development of Fc-engineered monoclonal antibodies with optimal Fc-effector function and improved clinical efficacy against COVID-19 disease.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Tratamento Farmacológico da COVID-19 , COVID-19/imunologia , Fragmentos Fc das Imunoglobulinas/imunologia , Fragmentos Fc das Imunoglobulinas/uso terapêutico , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/imunologia , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Anticorpos Neutralizantes/uso terapêutico , Cricetinae , Modelos Animais de Doenças , Feminino , Humanos , Fragmentos Fc das Imunoglobulinas/química , Fragmentos Fc das Imunoglobulinas/farmacologia , Imunoglobulina G/química , Imunoglobulina G/imunologia , Masculino , Camundongos , Profilaxia Pré-Exposição , Receptores de IgG/química , Receptores de IgG/imunologia , Resultado do TratamentoRESUMO
Rapidly emerging SARS-CoV-2 variants jeopardize antibody-based countermeasures. Although cell culture experiments have demonstrated a loss of potency of several anti-spike neutralizing antibodies against variant strains of SARS-CoV-21-3, the in vivo importance of these results remains uncertain. Here we report the in vitro and in vivo activity of a panel of monoclonal antibodies (mAbs), which correspond to many in advanced clinical development by Vir Biotechnology, AbbVie, AstraZeneca, Regeneron and Lilly, against SARS-CoV-2 variant viruses. Although some individual mAbs showed reduced or abrogated neutralizing activity in cell culture against B.1.351, B.1.1.28, B.1.617.1 and B.1.526 viruses with mutations at residue E484 of the spike protein, low prophylactic doses of mAb combinations protected against infection by many variants in K18-hACE2 transgenic mice, 129S2 immunocompetent mice and hamsters, without the emergence of resistance. Exceptions were LY-CoV555 monotherapy and LY-CoV555 and LY-CoV016 combination therapy, both of which lost all protective activity, and the combination of AbbVie 2B04 and 47D11, which showed a partial loss of activity. When administered after infection, higher doses of several mAb cocktails protected in vivo against viruses with a B.1.351 spike gene. Therefore, many-but not all-of the antibody products with Emergency Use Authorization should retain substantial efficacy against the prevailing variant strains of SARS-CoV-2.
Assuntos
Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais/farmacologia , Anticorpos Antivirais/uso terapêutico , COVID-19/virologia , Testes de Neutralização , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/imunologia , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/imunologia , COVID-19/genética , COVID-19/imunologia , COVID-19/prevenção & controle , Chlorocebus aethiops , Feminino , Humanos , Masculino , Mesocricetus/imunologia , Mesocricetus/virologia , Camundongos , Camundongos Transgênicos , Profilaxia Pós-Exposição , Profilaxia Pré-Exposição , SARS-CoV-2/genética , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Células VeroRESUMO
Tumor-targeting monoclonal antibodies are available for a number of cancer cell types (over)expressing the corresponding tumor antigens. Such antibodies can limit tumor progression by different mechanisms, including direct growth inhibition and immune-mediated mechanisms, in particular complement-dependent cytotoxicity, antibody-dependent cellular phagocytosis, and antibody-dependent cellular cytotoxicity (ADCC). ADCC can be mediated by various types of immune cells, including neutrophils, the most abundant leukocyte in circulation. Neutrophils express a number of Fc receptors, including Fcγ- and Fcα-receptors, and can therefore kill tumor cells opsonized with either IgG or IgA antibodies. In recent years, important insights have been obtained with respect to the mechanism(s) by which neutrophils engage and kill antibody-opsonized cancer cells and these findings are reviewed here. In addition, we consider a number of additional ways in which neutrophils may affect cancer progression, in particular by regulating adaptive anti-cancer immunity.
Assuntos
Neoplasias , Neutrófilos , Humanos , Citotoxicidade Celular Dependente de Anticorpos , Receptores Fc , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais/farmacologia , Receptores de IgG/metabolismoRESUMO
ABSTRACT: The CD161 inhibitory receptor is highly upregulated by tumor-infiltrating T cells in multiple human solid tumor types, and its ligand, CLEC2D, is expressed by both tumor cells and infiltrating myeloid cells. Here, we assessed the role of the CD161 receptor in hematological malignancies. Systematic analysis of CLEC2D expression using the Cancer Cell Line Encyclopedia revealed that CLEC2D messenger RNA was most abundant in hematological malignancies, including B-cell and T-cell lymphomas as well as lymphocytic and myelogenous leukemias. CLEC2D protein was detected by flow cytometry on a panel of cell lines representing a diverse set of hematological malignancies. We, therefore, used yeast display to generate a panel of high-affinity, fully human CD161 monoclonal antibodies (mAbs) that blocked CLEC2D binding. These mAbs were specific for CD161 and had a similar affinity for human and nonhuman primate CD161, a property relevant for clinical translation. A high-affinity CD161 mAb enhanced key aspects of T-cell function, including cytotoxicity, cytokine production, and proliferation, against B-cell lines originating from patients with acute lymphoblastic leukemia, diffuse large B-cell lymphoma, and Burkitt lymphoma. In humanized mouse models, this CD161 mAb enhanced T-cell-mediated immunity, resulting in a significant survival benefit. Single cell RNA-seq data demonstrated that CD161 mAb treatment enhanced expression of cytotoxicity genes by CD4 T cells as well as a tissue-residency program by CD4 and CD8 T cells that is associated with favorable survival outcomes in multiple human cancer types. These fully human mAbs, thus, represent potential immunotherapy agents for hematological malignancies.
Assuntos
Neoplasias Hematológicas , Neoplasias , Animais , Camundongos , Humanos , Linfócitos T CD4-Positivos , Imunidade Celular , Linfócitos T CD8-Positivos , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/terapia , Subfamília B de Receptores Semelhantes a Lectina de Células NK/genéticaRESUMO
ABSTRACT: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy. Current treatments, based on intensive chemotherapy regimens provide overall survival rates of â¼85% in children and <50% in adults, calling the search of new therapeutic options. We previously reported that targeting the T-cell receptor (TCR) in T-ALL with anti-CD3 (αCD3) monoclonal antibodies (mAbs) enforces a molecular program akin to thymic negative selection, a major developmental checkpoint in normal T-cell development; induces leukemic cell death; and impairs leukemia progression to ultimately improve host survival. However, αCD3 monotherapy resulted in relapse. To find out actionable targets able to re-enforce leukemic cells' vulnerability to αCD3 mAbs, including the clinically relevant teplizumab, we identified the molecular program induced by αCD3 mAbs in patient-derived xenografts derived from T-ALL cases. Using large-scale transcriptomic analysis, we found prominent expression of tumor necrosis factor α (TNFα), lymphotoxin α (LTα), and multiple components of the "TNFα via NF-κB signaling" pathway in anti-CD3-treated T-ALL. We show in vivo that etanercept, a sink for TNFα/LTα, enhances αCD3 antileukemic properties, indicating that TNF/TNF receptor (TNFR) survival pathways interferes with TCR-induced leukemic cell death. However, suppression of TNF-mediated survival and switch to TNFR-mediated cell death through inhibition of cellular inhibitor of apoptosis protein-1/2 (cIAP1/2) with the second mitochondrial-derived activator of caspases (SMAC) mimetic birinapant synergizes with αCD3 to impair leukemia expansion in a receptor-interacting serine/threonine-protein kinase 1-dependent manner and improve mice survival. Thus, our results advocate the use of either TNFα/LTα inhibitors, or birinapant/other SMAC mimetics to improve anti-CD3 immunotherapy in T-ALL.
Assuntos
Complexo CD3 , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Fator de Necrose Tumoral alfa , Humanos , Animais , Camundongos , Complexo CD3/imunologia , Complexo CD3/antagonistas & inibidores , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/terapia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/imunologia , Imunoterapia/métodos , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêuticoRESUMO
Cancer arises from malignant cells that exist in dynamic multilevel interactions with the host tissue. Cancer therapies aiming to directly kill cancer cells, including oncogene-targeted therapy and immune-checkpoint therapy that revives tumour-reactive cytotoxic T lymphocytes, are effective in some patients1,2, but acquired resistance frequently develops3,4. An alternative therapeutic strategy aims to rectify the host tissue pathology, including abnormalities in the vasculature that foster cancer progression5,6; however, neutralization of proangiogenic factors such as vascular endothelial growth factor A (VEGFA) has had limited clinical benefits7,8. Here, following the finding that transforming growth factor-ß (TGF-ß) suppresses T helper 2 (TH2)-cell-mediated cancer immunity9, we show that blocking TGF-ß signalling in CD4+ T cells remodels the tumour microenvironment and restrains cancer progression. In a mouse model of breast cancer resistant to immune-checkpoint or anti-VEGF therapies10,11, inducible genetic deletion of the TGF-ß receptor II (TGFBR2) in CD4+ T cells suppressed tumour growth. For pharmacological blockade, we engineered a bispecific receptor decoy by attaching the TGF-ß-neutralizing TGFBR2 extracellular domain to ibalizumab, a non-immunosuppressive CD4 antibody12,13, and named it CD4 TGF-ß Trap (4T-Trap). Compared with a non-targeted TGF-ß-Trap, 4T-Trap selectively inhibited TH cell TGF-ß signalling in tumour-draining lymph nodes, causing reorganization of tumour vasculature and cancer cell death, a process dependent on the TH2 cytokine interleukin-4 (IL-4). Notably, the 4T-Trap-induced tumour tissue hypoxia led to increased VEGFA expression. VEGF inhibition enhanced the starvation-triggered cancer cell death and amplified the antitumour effect of 4T-Trap. Thus, targeted TGF-ß signalling blockade in helper T cells elicits an effective tissue-level cancer defence response that can provide a basis for therapies directed towards the cancer environment.
Assuntos
Neoplasias da Mama/terapia , Imunoterapia , Transdução de Sinais/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/imunologia , Fator de Crescimento Transformador beta/antagonistas & inibidores , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Neoplasias da Mama/irrigação sanguínea , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Morte Celular/efeitos dos fármacos , Hipóxia Celular , Linhagem Celular Tumoral , Feminino , Células HEK293 , Humanos , Interleucina-4/imunologia , Linfonodos/citologia , Linfonodos/efeitos dos fármacos , Linfonodos/imunologia , Masculino , Camundongos , Receptor do Fator de Crescimento Transformador beta Tipo II/química , Receptor do Fator de Crescimento Transformador beta Tipo II/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Fator de Crescimento Transformador beta/imunologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly emerged coronavirus that is responsible for the current pandemic of coronavirus disease 2019 (COVID-19), which has resulted in more than 3.7 million infections and 260,000 deaths as of 6 May 20201,2. Vaccine and therapeutic discovery efforts are paramount to curb the pandemic spread of this zoonotic virus. The SARS-CoV-2 spike (S) glycoprotein promotes entry into host cells and is the main target of neutralizing antibodies. Here we describe several monoclonal antibodies that target the S glycoprotein of SARS-CoV-2, which we identified from memory B cells of an individual who was infected with severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003. One antibody (named S309) potently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2, by engaging the receptor-binding domain of the S glycoprotein. Using cryo-electron microscopy and binding assays, we show that S309 recognizes an epitope containing a glycan that is conserved within the Sarbecovirus subgenus, without competing with receptor attachment. Antibody cocktails that include S309 in combination with other antibodies that we identified further enhanced SARS-CoV-2 neutralization, and may limit the emergence of neutralization-escape mutants. These results pave the way for using S309 and antibody cocktails containing S309 for prophylaxis in individuals at a high risk of exposure or as a post-exposure therapy to limit or treat severe disease.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Betacoronavirus/imunologia , Reações Cruzadas/imunologia , Síndrome Respiratória Aguda Grave/imunologia , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Enzima de Conversão de Angiotensina 2 , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/farmacologia , Anticorpos Antivirais/química , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/farmacologia , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Linfócitos B/imunologia , Betacoronavirus/química , Betacoronavirus/efeitos dos fármacos , COVID-19 , Chlorocebus aethiops , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/terapia , Infecções por Coronavirus/virologia , Reações Cruzadas/efeitos dos fármacos , Microscopia Crioeletrônica , Epitopos de Linfócito B/química , Epitopos de Linfócito B/imunologia , Células HEK293 , Humanos , Evasão da Resposta Imune/imunologia , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/farmacologia , Memória Imunológica/imunologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Modelos Moleculares , Testes de Neutralização , Pandemias/prevenção & controle , Peptidil Dipeptidase A/química , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/imunologia , Pneumonia Viral/prevenção & controle , Pneumonia Viral/terapia , Pneumonia Viral/virologia , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/química , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/efeitos dos fármacos , SARS-CoV-2 , Síndrome Respiratória Aguda Grave/virologia , Glicoproteína da Espícula de Coronavírus/química , Células VeroRESUMO
Antibodies bind target molecules with exquisite specificity. The removal of these targets is mediated by the effector functions of antibodies. We reported earlier that the monoclonal antibody (mAb) 3F6 promotes opsonophagocytic killing of Staphylococcus aureus in blood and reduces bacterial replication in animals. Here, we generated mouse immunoglobulin G (mIgG) subclass variants and observed a hierarchy in protective efficacy 3F6-mIgG2a > 3F6-mIgG1 ≥ 3F6-mIgG2b >> 3F6-mIgG3 following bloodstream challenge of C57BL/6J mice. This hierarchy was not observed in BALB/cJ mice: All IgG subclasses conferred similar protection. IgG subclasses differ in their ability to activate complement and interact with Fcγ receptors (FcγR) on immune cells. 3F6-mIgG2a-dependent protection was lost in FcγR-deficient, but not in complement-deficient C57BL/6J animals. Measurements of the relative ratio of FcγRIV over complement receptor 3 (CR3) on neutrophils suggest the preferential expression of FcγRIV in C57BL/6 mice and of CR3 in BALB/cJ mice. To determine the physiological significance of these differing ratios, blocking antibodies against FcγRIV or CR3 were administered to animals before challenge. Correlating with the relative abundance of each receptor, 3F6-mIgG2a-dependent protection in C57BL/6J mice showed a greater reliance for FcγRIV while protection in BALB/cJ mice was only impaired upon neutralization of CR3. Thus, 3F6-based clearance of S. aureus in mice relies on a strain-specific contribution of variable FcγR- and complement-dependent pathways. We surmise that these variabilities are the result of genetic polymorphism(s) that may be encountered in other mammals including humans and may have clinical implications in predicting the efficacy of mAb-based therapies.
Assuntos
Imunoglobulina G , Staphylococcus aureus , Humanos , Camundongos , Animais , Staphylococcus aureus/metabolismo , Receptores de IgG/genética , Camundongos Endogâmicos C57BL , Anticorpos Monoclonais/farmacologia , Proteínas do Sistema Complemento , Mamíferos/metabolismoRESUMO
CD40 is a central costimulatory receptor implicated in productive antitumor immune responses across multiple cancers, including bladder cancer. Despite strong preclinical rationale, systemic administration of therapeutic agonistic antibodies targeting the CD40 pathway has demonstrated dose-limiting toxicities with minimal clinical activity, emphasizing an important need for optimized CD40-targeted approaches, including rational combination therapy strategies. Here, we describe a role for the endogenous IL-15 pathway in contributing to the therapeutic activity of CD40 agonism in orthotopic bladder tumors, with upregulation of transpresented IL-15/IL-15Rα surface complexes, particularly by cross-presenting conventional type 1 DCs (Dendritic Cells), and associated enrichment of activated CD8 T cells. In bladder cancer patient samples, we identify DCs as the primary source of IL-15, although they lack high levels of IL-15Rα at baseline. Using humanized immunocompetent orthotopic bladder tumor models, we demonstrate the ability to therapeutically augment this interaction through combined treatment with anti-CD40 agonist antibodies and exogenous IL-15, including the fully-human Fc-optimized antibody 2141-V11 currently in clinical development for the treatment of bladder cancer. Collectively, these data reveal an important role for IL-15 in mediating antitumor CD40 agonist responses in bladder cancer and provide key proof-of-concept for combined use of Fc-optimized anti-CD40 agonist antibodies and agents targeting the IL-15 pathway. These data support expansion of ongoing clinical studies evaluating anti-CD40 agonist antibodies and IL-15-based approaches to develop combinations of these promising therapeutics for the treatment of patients with bladder cancer.
Assuntos
Interleucina-15 , Neoplasias da Bexiga Urinária , Humanos , Neoplasias da Bexiga Urinária/tratamento farmacológico , Bexiga Urinária , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Terapia Combinada , Antígenos CD40 , Fragmentos Fc das ImunoglobulinasRESUMO
Antibodies play a central role in the immune defense against SARS-CoV-2. Emerging evidence has shown that nonneutralizing antibodies are important for immune defense through Fc-mediated effector functions. Antibody subclass is known to affect downstream Fc function. However, whether the antibody subclass plays a role in anti-SARS-CoV-2 immunity remains unclear. Here, we subclass-switched eight human IgG1 anti-spike monoclonal antibodies (mAbs) to the IgG3 subclass by exchanging their constant domains. The IgG3 mAbs exhibited altered avidities to the spike protein and more potent Fc-mediated phagocytosis and complement activation than their IgG1 counterparts. Moreover, combining mAbs into oligoclonal cocktails led to enhanced Fc- and complement receptor-mediated phagocytosis, superior to even the most potent single IgG3 mAb when compared at equivalent concentrations. Finally, in an in vivo model, we show that opsonic mAbs of both subclasses can be protective against a SARS-CoV-2 infection, despite the antibodies being nonneutralizing. Our results suggest that opsonic IgG3 oligoclonal cocktails are a promising idea to explore for therapy against SARS-CoV-2, its emerging variants, and potentially other viruses.