RESUMO
Anticytotoxic T lymphocyte-associated protein 4 (CTLA4) antibodies have shown potent antitumor activity, but systemic immune activation leads to severe immune-related adverse events, limiting clinical usage. We developed novel, conditionally active biologic (CAB) anti-CTLA4 antibodies that are active only in the acidic tumor microenvironment. In healthy tissue, this binding is reversibly inhibited by a novel mechanism using physiological chemicals as protein-associated chemical switches (PaCS). No enzymes or potentially immunogenic covalent modifications to the antibody are required for activation in the tumor. The novel anti-CTLA4 antibodies show similar efficacy in animal models compared to an analog of a marketed anti-CTLA4 biologic, but have markedly reduced toxicity in nonhuman primates (in combination with an anti-PD1 checkpoint inhibitor), indicating a widened therapeutic index (TI). The PaCS encompass mechanisms that are applicable to a wide array of antibody formats (e.g., ADC, bispecifics) and antigens. Examples shown here include antibodies to EpCAM, Her2, Nectin4, CD73, and CD3. Existing antibodies can be engineered readily to be made sensitive to PaCS, and the inhibitory activity can be optimized for each antigen's varying expression level and tissue distribution. PaCS can modulate diverse physiological molecular interactions and are applicable to various pathologic conditions, enabling differential CAB antibody activities in normal versus disease microenvironments.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais/farmacologia , Anticorpos Antineoplásicos/farmacologia , Antígeno B7-H1/antagonistas & inibidores , Antígeno CTLA-4/antagonistas & inibidores , Neoplasias do Colo/terapia , Imunoterapia/métodos , 5'-Nucleotidase/antagonistas & inibidores , 5'-Nucleotidase/genética , 5'-Nucleotidase/imunologia , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais Humanizados/química , Anticorpos Antineoplásicos/química , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Bicarbonatos/química , Complexo CD3/antagonistas & inibidores , Complexo CD3/genética , Complexo CD3/imunologia , Antígeno CTLA-4/genética , Antígeno CTLA-4/imunologia , Moléculas de Adesão Celular/antagonistas & inibidores , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/imunologia , Neoplasias do Colo/genética , Neoplasias do Colo/imunologia , Neoplasias do Colo/patologia , Molécula de Adesão da Célula Epitelial/antagonistas & inibidores , Molécula de Adesão da Célula Epitelial/genética , Molécula de Adesão da Célula Epitelial/imunologia , Proteínas Ligadas por GPI/antagonistas & inibidores , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Expressão Gênica , Humanos , Sulfeto de Hidrogênio/química , Concentração de Íons de Hidrogênio , Macaca fascicularis , Camundongos , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Engenharia de Proteínas/métodos , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/genética , Receptor ErbB-2/imunologia , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/patologia , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein that plays several roles in cancer biology. EpCAM is an attractive therapeutic target because of its expression in most solid tumors. However, targeting EpCAM has been challenging because it is also highly expressed in normal epithelial tissues. Initial attempts to develop EpCAM-specific T-cell engagers were unsuccessful due to severe cytokine release effects, as well as serious on-target, off-tumor drug-related toxicities. We developed novel, conditionally active biological (CAB) bispecific antibodies that bind to both EpCAM and CD3 in an acidic tumor microenvironment. In healthy tissues, binding to EpCAM and CD3 is greatly reduced by a novel, dual CAB selection, where each binding domain is independently blocked by the presence of physiological chemicals known as Protein-associated Chemical Switches (PaCS). The CAB anti-EpCAM T-cell engagers displayed the anticipated bispecific binding properties and mediated the potent lysis of EpCAM-positive cancer cell lines through the recruitment of T cells in the tumor microenvironment. Xenograft studies showed that the efficacy of CAB bispecific antibodies is similar to that of a non-CAB anti-EpCAM bispecific antibody, but they have markedly reduced toxicity in non-human primates, indicating an unprecedentedly widened therapeutic index of over 100-fold. These preclinical results indicate that the dual CAB bispecific antibody is potentially both a powerful and safe therapeutic platform and a promising T cell-engaging treatment for patients with EpCAM-expressing tumors.
Development of a novel conditionally active EpCAM-specific T-cell engager with enhanced safety and tolerability for treatment of solid tumors.
Assuntos
Anticorpos Biespecíficos , Produtos Biológicos , Neoplasias , Animais , Humanos , Molécula de Adesão da Célula Epitelial , Anticorpos Biespecíficos/farmacologia , Imunoterapia , Neoplasias/terapia , Microambiente TumoralRESUMO
The kinetics of drug-receptor interactions can profoundly influence in vivo and in vitro pharmacology. In vitro, the potencies of slowly associating agonists may be underestimated in assays capturing transient signaling events. When divergent receptor-mediated signaling pathways are evaluated using combinations of equilibrium and transient assays, potency differences driven by kinetics may be erroneously interpreted as biased signaling. In vivo, drugs with slow dissociation rates may display prolonged physiologic effects inconsistent with their pharmacokinetic profiles. We evaluated a panel of 5-hydroxytryptamine2B (5-HT2B) receptor agonists in kinetic radioligand binding assays and in transient, calcium flux assays, and inositol phosphate accumulation assays; two functional readouts emanating from Gαq-mediated activation of phospholipase C. In binding studies, ergot derivatives demonstrated slow receptor association and dissociation rates, resulting in significantly reduced potency in calcium assays relative to inositol phosphate accumulation assays. Ergot potencies for activation of extracellular signal-regulated kinases 1 and 2 were also highly time-dependent. A number of ergots produced wash-resistant 5-HT2B signaling that persisted for many hours without appreciable loss of potency, which was not explained simply by slow receptor-dissociation kinetics. Mechanistic studies indicated that persistent signaling originated from internalized or sequestered receptors. This study provides a mechanistic basis for the long durations of action in vivo and wash-resistant effects in ex vivo tissue models often observed for ergots. The 5-HT2B agonist activity of a number of ergot-derived therapeutics has been implicated in development of cardiac valvulopathy in man. The novel, sustained nature of ergot signaling reported here may represent an additional mechanism contributing to the valvulopathic potential of these compounds.
Assuntos
Receptor 5-HT2B de Serotonina/efeitos dos fármacos , Agonistas do Receptor de Serotonina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Antagonistas Adrenérgicos alfa/farmacologia , Anfetaminas/farmacocinética , Arrestinas/metabolismo , Cálcio/metabolismo , Relação Dose-Resposta a Droga , Alcaloides de Claviceps/farmacologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Células HEK293 , Humanos , Fenoxibenzamina/farmacologia , Fosforilação , Ensaio Radioligante , beta-ArrestinasRESUMO
BACKGROUND: Potassium (K+) channel inhibition has been identified in animal models as a potential target to increase pharyngeal dilator muscle activity and to treat OSA. However, these findings have not yet been translated to humans. RESEARCH QUESTION: Does a novel, potent, tandem of P domains in a weak inward rectifying K+ channel (TWIK)-related acid-sensitive K+ (TASK) 1/3 channel antagonist, BAY2586116, improve pharyngeal collapsibility in pigs and humans, and secondarily, what is the optimal dose and method of topical application? STUDY DESIGN AND METHODS: In the preclinical study, pharyngeal muscle activity and upper-airway collapsibility via transient negative pressure application was quantified in 13 anesthetized pigs during administration of placebo, 0.3 µg, 3 µg, and 30 µg nasal drops of BAY2586116. In the clinical study, 12 people with OSA instrumented with polysomnography equipment, an epiglottic pressure catheter, pneumotachograph, and nasal mask to monitor sleep and breathing performed up to four detailed upper airway sleep physiology studies. Participants received BAY2586116 (160 µg) or placebo nasal spray before sleep via a double-masked, randomized, crossover design. Most participants also returned for three additional overnight visits: (1) nasal drops (160 µg), (2) half-dose nasal spray (80 µg), and (3) direct endoscopic application (160 µg). The upper-airway critical closing pressure (Pcrit) during sleep was quantified at each visit. RESULTS: Consistent and sustained improvements in pharyngeal collapsibility to negative pressure were found with 3 and 30 µg of BAY2586116 vs placebo in pigs. Similarly, BAY2586116 improved pharyngeal collapsibility by an average of approximately 2 cm H2O vs placebo, regardless of topical application method and dose (P < .008, mixed model) in participants with OSA. INTERPRETATION: Acute topical application of BAY2586116 improves upper-airway collapsibility in anesthetized pigs and sleeping humans with OSA. These novel physiologic findings highlight the therapeutic potential to target potassium channel mechanisms to treat OSA. TRIAL REGISTRY: ClinicalTrials.gov; No.: NCT04236440; URL: www. CLINICALTRIALS: gov.
Assuntos
Apneia Obstrutiva do Sono , Humanos , Animais , Suínos , Apneia Obstrutiva do Sono/tratamento farmacológico , Sprays Nasais , Pressão Positiva Contínua nas Vias Aéreas , Faringe , Sono/fisiologiaRESUMO
Two series of fused tricyclic indoles were identified as potent and selective S1P(1) agonists. In vivo these agonists produced a significant reduction in circulating lymphocytes which translated into robust efficacy in several rodent models of autoimmune disease. Importantly, these agonists were devoid of any activity at the S1P(3) receptor in vitro, and correspondingly did not produce S1P(3) mediated bradycardia in telemeterized rat.
Assuntos
Fatores Imunológicos/química , Indóis/química , Receptores de Lisoesfingolipídeo/agonistas , Animais , Doenças Autoimunes/tratamento farmacológico , Modelos Animais de Doenças , Feminino , Humanos , Fatores Imunológicos/farmacocinética , Fatores Imunológicos/uso terapêutico , Indóis/farmacocinética , Indóis/uso terapêutico , Linfócitos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microssomos/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Lisoesfingolipídeo/metabolismo , Relação Estrutura-AtividadeRESUMO
S1P(1) receptor driven lymphopenia has proven utility in the treatment of an array of autoimmune disease states. As a part of our efforts to develop potent and selective S1P(1) receptor agonists, we have identified a novel chemical series of 4-oxo-4-(5-(5-phenyl-1,2,4-oxadiazol-3-yl)indolin-1-yl)butanoic acid S1P(1) receptor agonists.
Assuntos
Doenças Autoimunes/tratamento farmacológico , Butiratos/síntese química , Butiratos/farmacologia , Imunossupressores/síntese química , Imunossupressores/farmacologia , Indóis/síntese química , Indóis/farmacologia , Linfócitos/metabolismo , Proteínas do Tecido Nervoso/agonistas , Oxidiazóis/síntese química , Oxidiazóis/farmacologia , Proteínas de Ligação a RNA/agonistas , Animais , Butiratos/farmacocinética , Modelos Animais de Doenças , Cães , Relação Dose-Resposta a Droga , Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos , Haplorrinos , Ensaios de Triagem em Larga Escala , Humanos , Imunossupressores/farmacocinética , Indóis/farmacocinética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas do Tecido Nervoso/química , Oxidiazóis/farmacocinética , Proteínas de Ligação a RNA/química , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
We herein outline the design of a new series of agonists of the pancreatic and GI-expressed orphan G-protein coupled receptor GPR119, a target that has been of significant recent interest in the field of metabolism, starting from our prototypical agonist AR231453. A number of key parameters were improved first by incorporation of a pyrazolopyrimidine core to create a new structural series and secondly by the introduction of a piperidine ether group capped with a carbamate. Chronic treatment with one compound from the series, 3k, showed for the first time that blood glucose and glycated hemoglobin (HbA1c) levels could be significantly reduced in Zucker Diabetic Fatty (ZDF) rats over several weeks of dosing. As a result of these and other data described here, 3k (APD668, JNJ-28630368) was the first compound with this mechanism of action to be progressed into clinical development for the treatment of diabetes.
Assuntos
Glicemia/efeitos dos fármacos , Descoberta de Drogas , Hipoglicemiantes/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Animais , Compostos Bicíclicos Heterocíclicos com Pontes/síntese química , Compostos Bicíclicos Heterocíclicos com Pontes/química , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Modelos Animais de Doenças , Glucose/metabolismo , Hipoglicemiantes/síntese química , Hipoglicemiantes/química , Estrutura Molecular , Oxidiazóis/química , Oxidiazóis/farmacologia , Pirazóis/química , Pirazóis/farmacologia , Pirimidinas/química , Pirimidinas/farmacologia , Ratos , Ratos ZuckerRESUMO
G protein-coupled receptor 119 (GPR119) is largely restricted to pancreatic insulin-producing beta-cells and intestinal glucagon-like peptide-1-producing L-cells. Synthetic agonists of this receptor elicit glucose-dependent release of these endocrine factors, thereby enhancing glycemic control. Oleoylethanolamide also activates GPR119, but it remains unclear whether endogenous production of this lipid modulates GPR119 activity under normal or dysglycemic conditions. We show here that a relatively diverse set of lipid amides activate GPR119. Among these, the endovallinoid N-oleoyldopamine (OLDA) stimulated cAMP accumulation in GPR119-transfected cells as effectively as oleoylethanolamide and the previously described synthetic agonist AR231453. None of these lipid amides increased cAMP in control-transfected cells or in cells transfected with a number of other G protein-coupled receptors. OLDA stimulated both cAMP accumulation and insulin release in HIT-T15 cells, which express GPR119 endogenously, and in GPR119-transfected RIN-5F cells. Oral administration of OLDA to C57bl/6 mice elicited significant improvement in glucose tolerance, whereas GPR119-deficient mice were essentially unresponsive. OLDA also acutely elevated plasma gastric inhibitory peptide levels, a known hallmark of GPR119 activation. OLDA represents a possible paracrine modulator of GPR119 in pancreatic islets, where markers of dopamine synthesis correlated well with GPR119 expression. However, no such correlation was seen in the colon. Collectively, these studies indicate that multiple, distinct classes of lipid amides, acting via GPR119, may be important modulators of glucose homeostasis.
Assuntos
Glicemia/análise , Dopamina/análogos & derivados , Homeostase , Células Secretoras de Insulina/efeitos dos fármacos , Insulina/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Amidas/metabolismo , Amidas/farmacologia , Animais , Linhagem Celular , Cricetinae , Dopamina/metabolismo , Dopamina/farmacologia , Relação Dose-Resposta a Droga , Ácidos Graxos/metabolismo , Ácidos Graxos/farmacologia , Feminino , Homeostase/efeitos dos fármacos , Humanos , Secreção de Insulina , Células Secretoras de Insulina/enzimologia , Células Secretoras de Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Especificidade de Órgãos , Ratos , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/deficiência , Receptores Acoplados a Proteínas G/genéticaRESUMO
GPR119 is a rhodopsin-like GPCR expressed in pancreatic beta-cells and incretin releasing cells in the GI tract. As with incretins, GPR119 increases cAMP levels in these cell types, thus making it a highly attractive potential target for the treatment of diabetes. The discovery of the first reported potent agonist of GPR119, 2-fluoro-4-methanesulfonyl-phenyl)-{6-[4-(3-isopropyl-[1,2,4]oxadiazol-5-yl)-piperidin-1-yl]-5-nitro-pyrimidin-4-yl}-amine (8g, AR231453), is described starting from an initial inverse agonist screening hit. Compound 8g showed in vivo activity in rodents and was active in an oral glucose tolerance test in mice following oral administration.