RESUMO
Dedicator of cytokinesis 8 (DOCK8) is a guanine nucleotide exchange factor with an essential role in cytoskeletal rearrangement, cell migration, and survival of various immune cells. Interestingly, DOCK8-deficient mice are resistant to the development of experimental autoimmune encephalomyelitis (EAE). To understand if EAE resistance in these mice results from an alteration in dendritic cell (DC) functions, we generated mice with conditional deletion of DOCK8 in DCs and observed attenuated EAE in these mice compared with control mice. Additionally, we demonstrated that DOCK8 is important for the existence of splenic conventional DC2 and lymph node migratory DCs and further established that migratory DC, rather than resident DC, are essential for the generation and proliferation of pathogenic T cell populations upon immunization with myelin Ag in adjuvant. Therefore, our data suggest that limiting migratory DCs through DOCK8 deletion and possibly other mechanisms could limit the development of CNS autoimmunity.
Assuntos
Células Dendríticas/imunologia , Encefalomielite Autoimune Experimental/imunologia , Fatores de Troca do Nucleotídeo Guanina/imunologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Sensitization to fungi, such as the mold Aspergillus fumigatus, is increasingly becoming linked with asthma severity. We have previously shown that lung responses generated via the ß-glucan receptor Dectin-1 are required for lung defense during acute, invasive A. fumigatus infection. Unexpectedly, in an allergic model of chronic lung exposure to live A. fumigatus conidia, ß-glucan recognition via Dectin-1 led to the induction of multiple proallergic (Muc5ac, Clca3, CCL17, CCL22, and IL-33) and proinflammatory (IL-1ß and CXCL1) mediators that compromised lung function. Attenuated proallergic and proinflammatory responses in the absence of Dectin-1 were not associated with changes in Ido (IDO), Il12p35/Ebi3 (IL-35), IL-10, or TGF-ß levels. Assessment of Th responses demonstrated that purified lung CD4(+) T cells produced IL-4, IL-13, IFN-γ, and IL-17A, but not IL-22, in a Dectin-1-dependent manner. In contrast, we observed robust, Dectin-1-dependent IL-22 production by unfractionated lung digest cells. Intriguingly, the absence of IL-22 alone mimicked the attenuated proallergic and proinflammatory responses observed in the absence of Dectin-1, suggesting that Dectin-1-mediated IL-22 production potentiated responses that led to decrements in lung function. To this end, neutralization of IL-22 improved lung function in normal mice. Collectively, these results indicate that the ß-glucan receptor Dectin-1 contributes to lung inflammation and immunopathology associated with persistent fungal exposure via the production of IL-22.
Assuntos
Aspergillus fumigatus/imunologia , Glucanos/metabolismo , Interleucinas/fisiologia , Lectinas Tipo C/fisiologia , Receptores Imunológicos/fisiologia , Hipersensibilidade Respiratória/imunologia , Hipersensibilidade Respiratória/patologia , Animais , Células Cultivadas , Doença Crônica , Modelos Animais de Doenças , Interleucinas/deficiência , Interleucinas/genética , Lectinas Tipo C/deficiência , Lectinas Tipo C/genética , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/patologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hipersensibilidade Respiratória/microbiologia , Interleucina 22RESUMO
Increasing use of covalent and noncovalent inhibitors of Bruton's tyrosine kinase (BTK) has elucidated a series of acquired drug-resistant BTK mutations in patients with B cell malignancies. Here we identify inhibitor resistance mutations in BTK with distinct enzymatic activities, including some that impair BTK enzymatic activity while imparting novel protein-protein interactions that sustain B cell receptor (BCR) signaling. Furthermore, we describe a clinical-stage BTK and IKZF1/3 degrader, NX-2127, that can bind and proteasomally degrade each mutant BTK proteoform, resulting in potent blockade of BCR signaling. Treatment of chronic lymphocytic leukemia with NX-2127 achieves >80% degradation of BTK in patients and demonstrates proof-of-concept therapeutic benefit. These data reveal an oncogenic scaffold function of mutant BTK that confers resistance across clinically approved BTK inhibitors but is overcome by BTK degradation in patients.
Assuntos
Tirosina Quinase da Agamaglobulinemia , Resistencia a Medicamentos Antineoplásicos , Fator de Transcrição Ikaros , Leucemia Linfocítica Crônica de Células B , Inibidores de Proteínas Quinases , Proteólise , Humanos , Tirosina Quinase da Agamaglobulinemia/genética , Tirosina Quinase da Agamaglobulinemia/metabolismo , Fator de Transcrição Ikaros/metabolismo , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/genética , Mutação , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Transdução de Sinais , Proteólise/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacosRESUMO
Chlorine (Cl2) is a highly irritating and reactive gas with potential occupational and environmental hazards. Acute exposure to Cl2 induces severe epithelial damage, airway hyperreactivity, impaired alveolar fluid clearance, and pulmonary edema in the presence of heightened inflammation and significant neutrophil accumulation in the lungs. Herein, we investigated whether Cl2 exposure affected the lung antimicrobial immune response leading to increased susceptibility to opportunistic infections. Mice exposed to Cl2 and challenged intratracheally 24 h thereafter with the opportunistic mold Aspergillus fumigatus demonstrated an >500-fold increase in A. fumigatus lung burden 72 h postchallenge compared with A. fumigatus mice exposed to room air. Cl2-exposed A. fumigatus challenged mice also demonstrated significantly higher lung resistance following methacholine challenge and increased levels of plasma proteins (albumin and IgG) in the bronchoalveolar lavage fluid. Despite enhanced recruitment of inflammatory cells to the lungs of Cl2-exposed A. fumigatus challenged mice, these cells (>60% of which were neutrophils) demonstrated a profound impairment in generating superoxide. Significantly higher A. fumigatus burden in the lungs of Cl2 exposed mice correlated with enhanced production of IL-6, TNF-α, CXCL1, CCL2, and CCL3. Surprisingly, however, Cl2-exposed A. fumigatus challenged mice had a specific impairment in the production of IL-17A and IL-22 in the lungs compared with mice exposed to room air and challenged with A. fumigatus. In summary, our results indicate that Cl2 exposure markedly impairs the antimicrobial activity and inflammatory reactivity of myeloid cells in the lung leading to increased susceptibility to opportunistic pathogens.
Assuntos
Aspergilose/etiologia , Cloro/toxicidade , Pneumopatias Fúngicas/etiologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Quimiocinas/metabolismo , Citocinas/metabolismo , Suscetibilidade a Doenças , Interleucina-17/biossíntese , Interleucina-6/biossíntese , Interleucinas/biossíntese , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/fisiologia , Pneumopatias Fúngicas/imunologia , Pneumopatias Fúngicas/microbiologia , Masculino , Camundongos , Superóxidos/metabolismo , Interleucina 22RESUMO
We have previously reported that mice deficient in the beta-glucan receptor Dectin-1 displayed increased susceptibility to Aspergillus fumigatus lung infection in the presence of lower interleukin 23 (IL-23) and IL-17A production in the lungs and have reported a role for IL-17A in lung defense. As IL-23 is also thought to control the production of IL-22, we examined the role of Dectin-1 in IL-22 production, as well as the role of IL-22 in innate host defense against A. fumigatus. Here, we show that Dectin-1-deficient mice demonstrated significantly reduced levels of IL-22 in the lungs early after A. fumigatus challenge. Culturing cells from enzymatic lung digests ex vivo further demonstrated Dectin-1-dependent IL-22 production. IL-22 production was additionally found to be independent of IL-1ß, IL-6, or IL-18 but required IL-23. The addition of recombinant IL-23 augmented IL-22 production in wild-type (WT) lung cells and rescued IL-22 production by lung cells from Dectin-1-deficient mice. In vivo neutralization of IL-22 in the lungs of WT mice resulted in impaired A. fumigatus lung clearance. Moreover, mice deficient in IL-22 also demonstrated a higher lung fungal burden after A. fumigatus challenge in the presence of impaired IL-1α, tumor necrosis factor alpha (TNF-α), CCL3/MIP-1α, and CCL4/MIP-1ß production and lower neutrophil recruitment, yet intact IL-17A production. We further show that lung lavage fluid collected from both A. fumigatus-challenged Dectin-1-deficient and IL-22-deficient mice had compromised anti-fungal activity against A. fumigatus in vitro. Although lipocalin 2 production was observed to be Dectin-1 and IL-22 dependent, lipocalin 2-deficient mice did not demonstrate impaired A. fumigatus clearance. Moreover, lung S100a8, S100a9, and Reg3g mRNA expression was not lower in either Dectin-1-deficient or IL-22-deficient mice. Collectively, our results indicate that early innate lung defense against A. fumigatus is mediated by Dectin-1-dependent IL-22 production.
Assuntos
Aspergillus fumigatus/imunologia , Interleucinas/imunologia , Lectinas Tipo C/metabolismo , Pulmão/imunologia , Aspergilose Pulmonar/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Células Cultivadas , Contagem de Colônia Microbiana , Lectinas Tipo C/deficiência , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/imunologia , Interleucina 22RESUMO
We have previously reported that compromised interleukin 17A (IL-17A) production in the lungs increased susceptibility to infection with the invasive fungal pathogen Aspergillus fumigatus. Here we have shown that culturing lung cells from A. fumigatus-challenged mice ex vivo demonstrated Dectin-1-dependent IL-17A production. In this system, neutralization of IL-23 but not IL-6, IL-1ß, or IL-18 resulted in attenuated IL-17A production. Il23 mRNA expression was found to be lower in lung cells from A. fumigatus-challenged Dectin-1-deficient mice, whereas bone marrow-derived dendritic cells from Dectin-1-deficient mice failed to produce IL-23 in response to A. fumigatus in vitro. Addition of recombinant IL-23 augmented IL-17A production by wild-type (WT) and Dectin-1-deficient lung cells, although the addition of IL-6 or IL-1ß did not augment the effect of IL-23. Intracellular cytokine staining of lung cells revealed lower levels of CD11b(+) IL-17A(+) and Ly-6G(+) IL-17A(+) cells in A. fumigatus-challenged Dectin-1-deficient mice. Ly-6G(+) neutrophils purified from the lungs of A. fumigatus-challenged Dectin-1-deficient mice displayed lower Il17a mRNA expression but surprisingly had intact Rorc and Rora mRNA expression. We further demonstrated that Ly-6G(+) neutrophils required the presence of myeloid cells for IL-17A production. Finally, upon in vitro stimulation with A. fumigatus, thioglycolate-elicited peritoneal neutrophils were positive for intracellular IL-17A expression and produced IL-17A in a Dectin-1- and IL-23-dependent manner. In summary, Dectin-1-dependent IL-17A production in the lungs during invasive fungal infection is mediated in part by CD11b(+) Ly-6G(+) neutrophils in an IL-23-dependent manner.
Assuntos
Aspergillus fumigatus/patogenicidade , Interleucina-17/biossíntese , Interleucina-23/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neutrófilos/metabolismo , Aspergilose Pulmonar/imunologia , Animais , Aspergillus fumigatus/imunologia , Células Cultivadas , Interleucina-23/deficiência , Interleucina-23/genética , Lectinas Tipo C , Pulmão/citologia , Pulmão/imunologia , Pulmão/metabolismo , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Neutrófilos/imunologia , Aspergilose Pulmonar/microbiologiaRESUMO
This was a randomized, single-blind, single-dose, 3-arm parallel-group study. Healthy subjects were randomized to receive ABP 710 (n = 50) or infliximab reference product (RP) sourced from the United States (infliximab US; n = 50) or the European Union (infliximab EU; n = 50) 5 mg/kg intravenously over 2 hours. The primary endpoint was area under the serum concentration-time curve from time 0 extrapolated to infinity (AUCinf ) for the comparison of ABP 710 to infliximab US and infliximab EU. Secondary endpoints included safety, tolerability, and immunogenicity. AUCinf was similar across the 3 groups, showing similarity of ABP 710 to infliximab RP as well as similarity of infliximab US with infliximab EU. Geometric mean ratio of AUCinf was 0.89 between ABP 710 and infliximab US, 1.00 between ABP 710 and infliximab EU, and 1.11 between infliximab US and infliximab EU. All 90% confidence intervals of the geometric mean ratios were fully contained within the prespecified standard pharmacokinetic equivalence criteria range of 0.80 to 1.25. Treatment-related adverse events were mild to moderate and reported for 83.7%, 86.0%, and 83.7% of subjects in the ABP 710, infliximab US, and infliximab EU treatment groups, respectively; incidence of antidrug antibody rates observed across the 3 groups were similar. Results of this study demonstrated pharmacokinetic similarity of ABP 710 with infliximab RP following a single 5-mg/kg intravenous injection. The safety and tolerability of ABP 710 and infliximab RP were comparable. These results add to the totality of evidence providing further support that the proposed biosimilar ABP 710 is similar to infliximab RP. (Trial ID: ACTRN12614000903684.).
Assuntos
Anticorpos Monoclonais/farmacocinética , Antirreumáticos/farmacocinética , Infliximab/farmacocinética , Inibidores do Fator de Necrose Tumoral/farmacocinética , Administração Intravenosa , Adulto , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/sangue , Antirreumáticos/administração & dosagem , Antirreumáticos/efeitos adversos , Antirreumáticos/sangue , Área Sob a Curva , Austrália , Medicamentos Biossimilares/administração & dosagem , Medicamentos Biossimilares/efeitos adversos , Medicamentos Biossimilares/sangue , Medicamentos Biossimilares/farmacocinética , União Europeia , Feminino , Voluntários Saudáveis , Humanos , Infliximab/administração & dosagem , Infliximab/efeitos adversos , Infliximab/sangue , Masculino , Pessoa de Meia-Idade , Segurança , Método Simples-Cego , Inibidores do Fator de Necrose Tumoral/administração & dosagem , Inibidores do Fator de Necrose Tumoral/efeitos adversos , Inibidores do Fator de Necrose Tumoral/sangue , Estados UnidosRESUMO
ABP 798 is a proposed biosimilar to rituximab reference product (RP), an anti-CD20 monoclonal antibody. Pharmacokinetics (PK), pharmacodynamics (PD), and safety results from the comparative clinical study that evaluated the PK, PD, safety, efficacy, and immunogenicity of ABP 798 versus rituximab RP are presented here. Subjects with moderate to severe rheumatoid arthritis (RA) received 2 doses of ABP 798, United States-sourced RP (rituximab US) or European Union-sourced RP (rituximab EU), each consisting of two 1000-mg infusions 2 weeks apart. For the second dose (week 24), ABP 798- and rituximab EU-treated subjects received the same treatment; rituximab US-treated subjects transitioned to ABP 798. End points included area under the serum concentration-time curve from time 0 extrapolated to infinity and maximum observed serum concentration following the second infusion of the first dose (PK) and percentage of subjects with complete CD19+ cell depletion days 1-33 (PD). Primary analysis established PK similarity between ABP 798 and rituximab RP based on 90% confidence intervals of the adjusted geometric mean ratios being within a prespecified equivalence margin of 0.8 and 1.25. Complete CD19+ B-cell depletion on day 3 among groups confirmed PD similarity. These findings demonstrated PK/PD similarity between ABP 798 and rituximab RP in subjects with moderate to severe RA.
Assuntos
Antirreumáticos/farmacocinética , Artrite Reumatoide/tratamento farmacológico , Medicamentos Biossimilares/farmacocinética , Liases de Carbono-Enxofre/farmacocinética , Rituximab/farmacocinética , Adulto , Idoso , Antineoplásicos Imunológicos/administração & dosagem , Antineoplásicos Imunológicos/sangue , Antineoplásicos Imunológicos/farmacocinética , Antineoplásicos Imunológicos/farmacologia , Antirreumáticos/administração & dosagem , Antirreumáticos/sangue , Antirreumáticos/farmacologia , Área Sob a Curva , Medicamentos Biossimilares/administração & dosagem , Medicamentos Biossimilares/sangue , Medicamentos Biossimilares/farmacologia , Liases de Carbono-Enxofre/administração & dosagem , Liases de Carbono-Enxofre/sangue , Liases de Carbono-Enxofre/farmacologia , Método Duplo-Cego , Feminino , Humanos , Infusões Intravenosas , Masculino , Pessoa de Meia-Idade , Rituximab/administração & dosagem , Rituximab/sangue , Rituximab/farmacologia , Segurança , Índice de Gravidade de Doença , Equivalência Terapêutica , Resultado do TratamentoRESUMO
Cancer vaccines using synthetic long peptides (SLP) targeting tumor antigens have been tested in the clinic but the outcomes have been unimpressive, perhaps because these peptides elicit predominantly CD4+ T cell responses. We hypothesized that enhanced delivery of peptide antigens to, and uptake in, secondary lymphoid tissues should elicit more robust CD8+ and CD4+ T cell responses and improved anti-tumor responses. Here, we have designed SLP-containing cationic lipoplexes (SLP-Lpx) that improve delivery of peptides to myeloid cells in the spleen and lymphatics. Using the G12D KRAS mutations as neoantigens, we found that vaccination of mice with naked synthetic peptides harboring the G12D mutation with CpG adjuvant stimulated mainly CD4+ T cell responses with limited tumor growth inhibition. On the other hand, immunization with SLP-Lpx stimulated both CD4+ and CD8+ T cells and suppressed tumor growth in a CD8+ T cell-dependent manner. Combination of the SLP-Lpx vaccines with a checkpoint inhibitor led to profound growth suppression of established tumors. These studies suggest that preferential targeting of peptides derived from neoantigens to the spleen via lipoplexes elicits potent CD4+ and CD8+ T cell responses that inhibit tumor growth.