RESUMEN
Targeted therapies that suppress B cell receptor (BCR) signaling have emerged as promising agents in autoimmune disease and B cell malignancies. Bruton's tyrosine kinase (Btk) plays a crucial role in B cell development and activation through the BCR signaling pathway and represents a new target for diseases characterized by inappropriate B cell activity. N-(3-(5-fluoro-2-(4-(2-methoxyethoxy)phenylamino)pyrimidin-4-ylamino)phenyl)acrylamide (CC-292) is a highly selective, covalent Btk inhibitor and a sensitive and quantitative assay that measures CC-292-Btk engagement has been developed. This translational pharmacodynamic assay has accompanied CC-292 through each step of drug discovery and development. These studies demonstrate the quantity of Btk bound by CC-292 correlates with the efficacy of CC-292 in vitro and in the collagen-induced arthritis model of autoimmune disease. Recently, CC-292 has entered human clinical trials with a trial design that has provided rapid insight into safety, pharmacokinetics, and pharmacodynamics. This first-in-human healthy volunteer trial has demonstrated that a single oral dose of 2 mg/kg CC-292 consistently engaged all circulating Btk protein and provides the basis for rational dose selection in future clinical trials. This targeted covalent drug design approach has enabled the discovery and early clinical development of CC-292 and has provided support for Btk as a valuable drug target for B-cell mediated disorders.
Asunto(s)
Acrilamidas/farmacología , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Pirimidinas/farmacología , Acrilamidas/farmacocinética , Acrilamidas/uso terapéutico , Agammaglobulinemia Tirosina Quinasa , Animales , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/inmunología , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/inmunología , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Método Doble Ciego , Humanos , Ratones , Pirimidinas/farmacocinética , Pirimidinas/uso terapéutico , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de SeñalRESUMEN
Designing selective inhibitors of proteases has proven problematic, in part because pharmacophores that confer potency exploit the conserved catalytic apparatus. We developed a fundamentally different approach by designing irreversible inhibitors that target noncatalytic cysteines that are structurally unique to a target in a protein family. We have successfully applied this approach to the important therapeutic target HCV protease, which has broad implications for the design of other selective protease inhibitors.
Asunto(s)
Inhibidores de Cisteína Proteinasa/uso terapéutico , Cisteína/antagonistas & inhibidores , Diseño de Fármacos , Oligopéptidos/uso terapéutico , Biocatálisis , Bioquímica/métodos , Cristalografía por Rayos X , Cisteína/metabolismo , Inhibidores de Cisteína Proteinasa/química , Inhibidores de Cisteína Proteinasa/farmacología , Hepacivirus/efectos de los fármacos , Hepacivirus/enzimología , Hepacivirus/crecimiento & desarrollo , Oligopéptidos/química , Oligopéptidos/farmacología , Virología/métodosRESUMEN
PURPOSE: While immune checkpoint inhibitors such as anti-PD-L1 are rapidly becoming the standard of care in the treatment of many cancers, only a subset of treated patients have long-term responses. IL12 promotes antitumor immunity in mouse models; however, systemic recombinant IL12 had significant toxicity and limited efficacy in early clinical trials. EXPERIMENTAL DESIGN: We therefore designed a novel intratumoral IL12 mRNA therapy to promote local IL12 tumor production while mitigating systemic effects. RESULTS: A single intratumoral dose of mouse (m)IL12 mRNA induced IFNγ and CD8+ T-cell-dependent tumor regression in multiple syngeneic mouse models, and animals with a complete response demonstrated immunity to rechallenge. Antitumor activity of mIL12 mRNA did not require NK and NKT cells. mIL12 mRNA antitumor activity correlated with TH1 tumor microenvironment (TME) transformation. In a PD-L1 blockade monotherapy-resistant model, antitumor immunity induced by mIL12 mRNA was enhanced by anti-PD-L1. mIL12 mRNA also drove regression of uninjected distal lesions, and anti-PD-L1 potentiated this response. Importantly, intratumoral delivery of mRNA encoding membrane-tethered mIL12 also drove rejection of uninjected lesions with very limited circulating IL12p70, supporting the hypothesis that local IL12 could induce a systemic antitumor immune response against distal lesions. Furthermore, in ex vivo patient tumor slice cultures, human IL12 mRNA (MEDI1191) induced dose-dependent IL12 production, downstream IFNγ expression and TH1 gene expression. CONCLUSIONS: These data demonstrate the potential for intratumorally delivered IL12 mRNA to promote TH1 TME transformation and robust antitumor immunity.See related commentary by Cirella et al., p. 6080.
Asunto(s)
Neoplasias Colorrectales/prevención & control , Interleucina-12/administración & dosificación , Linfocitos Infiltrantes de Tumor/inmunología , Melanoma/prevención & control , ARN Mensajero/administración & dosificación , Células TH1/inmunología , Microambiente Tumoral/inmunología , Animales , Anticuerpos Monoclonales/farmacología , Apoptosis , Antígeno B7-H1/antagonistas & inhibidores , Linfocitos T CD8-positivos , Proliferación Celular , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/inmunología , Neoplasias Colorrectales/patología , Resistencia a Antineoplásicos , Femenino , Humanos , Interleucina-12/genética , Melanoma/genética , Melanoma/inmunología , Melanoma/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Desnudos , Ratones SCID , ARN Mensajero/genética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Many solid cancers contain dysfunctional immune microenvironments. Immune system modulators that initiate responses to foreign pathogens could be promising candidates for reigniting productive responses toward tumors. Interleukin-1 (IL-1) and IL-12 cytokine family members cooperate at barrier tissues after microbial invasion, in human inflammatory diseases, and in antitumoral immunity. IL-36γ, in classic alarmin fashion, acts in damaged tissues, whereas IL-23 centrally coordinates immune responses to danger signals. In this study, direct intratumoral delivery of messenger RNAs (mRNAs) encoding these cytokines produced robust anticancer responses in a broad range of tumor microenvironments. The addition of mRNA encoding the T cell costimulator OX40L increased complete response rates in treated and untreated distal tumors compared to the cytokine mRNAs alone. Mice exhibiting complete responses were subsequently protected from tumor rechallenge. Treatments with these mRNA mixtures induced downstream cytokine and chemokine expression, and also activated multiple dendritic cell (DC) and T cell types. Consistent with this, efficacy was dependent on Batf3-dependent cross-presenting DCs and cytotoxic CD8+ T cells. IL-23/IL-36γ/OX40L triplet mRNA mixture triggered substantial immune cell recruitment into tumors, enabling effective tumor destruction irrespective of previous tumoral immune infiltrates. Last, combining triplet mRNA with checkpoint blockade led to efficacy in models otherwise resistant to systemic immune checkpoint inhibition. Human cell studies showed similar cytokine responses to the individual components of this mRNA mixture, suggesting translatability of immunomodulatory activity to human patients.
Asunto(s)
Inmunidad , Interleucina-1/genética , Interleucina-23/genética , Neoplasias/inmunología , Ligando OX40/genética , ARN Mensajero/administración & dosificación , Animales , Proliferación Celular , Modelos Animales de Enfermedad , Humanos , Inflamación/patología , Interleucina-1/metabolismo , Interleucina-23/metabolismo , Ganglios Linfáticos/patología , Activación de Linfocitos/inmunología , Ratones , Ligando OX40/metabolismo , Distribución Tisular , Microambiente Tumoral/inmunologíaRESUMEN
PURPOSE: Fumagillin and related compounds have potent antiproliferative activity through inhibition of methionine aminopeptidase-2 (MetAP-2). It has recently been reported that MetAP-2 is highly expressed in germinal center B cells and germinal center-derived non-Hodgkin's lymphomas (NHL), suggesting an important role for MetAP-2 in proliferating B cells. Therefore, we determined the importance of MetAP-2 in normal and transformed germinal center B cells by evaluating the effects of MetAP-2 inhibition on the form and function of germinal centers and germinal center-derived NHL cells. EXPERIMENTAL DESIGN: To examine the activity of PPI-2458 on germinal center morphology, spleen sections from cynomolgus monkeys treated with oral PPI-2458 were analyzed. Antiproliferative activity of PPI-2458 was assessed on germinal center-derived NHL lines in culture. A MetAP-2 pharmacodynamic assay was used to determine cellular MetAP-2 inhibition following PPI-2458 treatment. Finally, inhibition of MetAP-2 and proliferation by PPI-2458 was examined in the human SR NHL line in culture and in implanted xenografts. RESULTS: Oral PPI-2458 caused a reduction in germinal center size and number in lymphoid tissues from treated animals. PPI-2458 potently inhibited growth (GI(50) = 0.2-1.9 nmol/L) of several NHL lines in a manner that correlated with MetAP-2 inhibition. Moreover, orally administered PPI-2458 significantly inhibited SR tumor growth, which correlated with inhibition of tumor MetAP-2 (>85% at 100 mg/kg) in mice. CONCLUSIONS: These results show the potent antiproliferative activity of PPI-2458 on NHL lines in vitro and oral antitumor activity in vivo and suggest the therapeutic potential of PPI-2458 as a novel agent for treatment of NHL should be evaluated in the clinical setting.
Asunto(s)
Aminopeptidasas/antagonistas & inhibidores , Proliferación Celular/efectos de los fármacos , Compuestos Epoxi/farmacología , Linfoma no Hodgkin/tratamiento farmacológico , Metaloendopeptidasas/antagonistas & inhibidores , Valina/análogos & derivados , Aminopeptidasas/metabolismo , Animales , Linfocitos B/efectos de los fármacos , Linfocitos B/patología , Western Blotting , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Compuestos Epoxi/uso terapéutico , Femenino , Centro Germinal/efectos de los fármacos , Centro Germinal/patología , Humanos , Recuento de Linfocitos , Linfoma de Células B/tratamiento farmacológico , Linfoma de Células B/metabolismo , Linfoma de Células B/patología , Linfoma no Hodgkin/metabolismo , Linfoma no Hodgkin/patología , Macaca fascicularis , Metaloendopeptidasas/metabolismo , Ratones , Ratones SCID , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Factores de Tiempo , Valina/farmacología , Valina/uso terapéutico , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Over the past few decades, melanoma has shown the fastest growing incidence rate of all cancers. This malignancy is clinically defined by its potential to rapidly metastasize, and advanced metastatic melanomas are highly resistant to existing therapeutic regimens. Here, we report that PPI-2458, a novel, orally active agent of the fumagillin class of irreversible methionine aminopeptidase-2 (MetAP-2) inhibitors, potently inhibited the proliferation of B16F10 melanoma cells in vitro, with a growth inhibitory concentration 50% (GI50) of 0.2 nM. B16F10 growth inhibition was correlated with the inhibition of MetAP-2 enzyme, in a dose-dependent fashion, as determined by a pharmacodynamic assay, which measures the amount of uninhibited MetAP-2 following PPI-2458 treatment. Prolonged exposure of B16F10 cells to PPI-2458 at concentrations of up to 1 microM, 5,000-fold above the GI50, did not alter their sensitivity to PPI-2458 growth inhibition and no drug resistance was observed. Moreover, prolonged exposure to this agent induced melanogenesis, concomitant with the elevated expression of the melanocyte-specific enzymes tyrosinase and tyrosinase-related proteins (TRP) 1 and 2, a morphological feature associated with differentiated melanocytes. PPI-2458, when administered orally (p.o.), significantly inhibited B16F10 tumor growth in mice in a dose-dependent fashion, with a maximum inhibition of 62% at 100 mg/kg. This growth inhibition was directly correlated to the amount of irreversibly inhibited MetAP-2 (80% at 100 mg/kg PPI-2458) in tumor tissue. These data demonstrate that PPI-2458 has potent antiproliferative activity against B16F10 cells in vitro and in vivo, and that both activities are directly correlated with levels of MetAP-2 enzyme inhibition. This antiproliferative activity, coupled with additional observations from studies in vitro (absence of detectable resistance to PPI-2458 and induction of morphological features consistent with differentiated melanocytes), provides a rationale for assessing the therapeutic potential of PPI-2458 in the treatment of melanoma.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Compuestos Epoxi/farmacología , Melanoma Experimental/prevención & control , Valina/análogos & derivados , Administración Oral , Aminopeptidasas/antagonistas & inhibidores , Aminopeptidasas/metabolismo , Animales , Western Blotting , Línea Celular , Línea Celular Tumoral , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Relación Dosis-Respuesta a Droga , Compuestos Epoxi/administración & dosificación , Compuestos Epoxi/uso terapéutico , Glicoproteínas/antagonistas & inhibidores , Glicoproteínas/metabolismo , Humanos , Masculino , Melaninas/metabolismo , Melanoma Experimental/metabolismo , Melanoma Experimental/patología , Metionil Aminopeptidasas , Ratones , Ratones Endogámicos C57BL , Valina/administración & dosificación , Valina/farmacología , Valina/uso terapéuticoRESUMEN
Patients with non-small cell lung carcinoma (NSCLC) with activating mutations in epidermal growth factor receptor (EGFR) initially respond well to the EGFR inhibitors erlotinib and gefitinib. However, all patients relapse because of the emergence of drug-resistant mutations, with T790M mutations accounting for approximately 60% of all resistance. Second-generation irreversible EGFR inhibitors are effective against T790M mutations in vitro, but retain affinity for wild-type EGFR (EGFR(WT)). These inhibitors have not provided compelling clinical benefit in T790M-positive patients, apparently because of dose-limiting toxicities associated with inhibition of EGFR(WT). Thus, there is an urgent clinical need for therapeutics that overcome T790M drug resistance while sparing EGFR(WT). Here, we describe a lead optimization program that led to the discovery of four potent irreversible 2,4-diaminopyrimidine compounds that are EGFR mutant (EGFR(mut)) selective and have been designed to have low affinity for EGFR(WT). Pharmacokinetic and pharmacodynamic studies in H1975 tumor-bearing mice showed that exposure was dose proportional resulting in dose-dependent EGFR modulation. Importantly, evaluation of normal lung tissue from the same animals showed no inhibition of EGFR(WT). Of all the compounds tested, compound 3 displayed the best efficacy in EGFR(L858R/T790M)-driven tumors. Compound 3, now renamed CO-1686, is currently in a phase I/II clinical trial in patients with EGFR(mut)-advanced NSCLC that have received prior EGFR-directed therapy.
Asunto(s)
4-Aminopiridina/análogos & derivados , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Receptores ErbB/metabolismo , Recurrencia Local de Neoplasia/tratamiento farmacológico , 4-Aminopiridina/administración & dosificación , Animales , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ensayos Clínicos como Asunto , Resistencia a Antineoplásicos/genética , Receptores ErbB/antagonistas & inhibidores , Humanos , Técnicas In Vitro , Ratones , Mutación , Recurrencia Local de Neoplasia/metabolismo , Recurrencia Local de Neoplasia/patología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
PI3Kα has been identified as an oncogene in human tumors. By use of rational drug design, a targeted covalent inhibitor 3 (CNX-1351) was created that potently and specifically inhibits PI3Kα. We demonstrate, using mass spectrometry and X-ray crystallography, that the selective inhibitor covalently modifies PI3Kα on cysteine 862 (C862), an amino acid unique to the α isoform, and that PI3Kß, -γ, and -δ are not covalently modified. 3 is able to potently (EC(50) < 100 nM) and specifically inhibit signaling in PI3Kα-dependent cancer cell lines, and this leads to a potent antiproliferative effect (GI(50) < 100 nM). A covalent probe, 8 (CNX-1220), which selectively bonds to PI3Kα, was used to investigate the duration of occupancy of 3 with PI3Kα in vivo. This is the first report of a PI3Kα-selective inhibitor, and these data demonstrate the biological impact of selectively targeting PI3Kα.
Asunto(s)
Descubrimiento de Drogas , Isoenzimas/antagonistas & inhibidores , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacología , Animales , Masculino , Espectrometría de Masas , Ratones , Ratones Endogámicos C57BL , Resonancia Magnética Nuclear Biomolecular , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/química , Transducción de SeñalRESUMEN
UNLABELLED: Patients with non-small cell lung cancer (NSCLC) with activating EGF receptor (EGFR) mutations initially respond to first-generation reversible EGFR tyrosine kinase inhibitors. However, clinical efficacy is limited by acquired resistance, frequently driven by the EGFR(T790M) mutation. CO-1686 is a novel, irreversible, and orally delivered kinase inhibitor that specifically targets the mutant forms of EGFR, including T790M, while exhibiting minimal activity toward the wild-type (WT) receptor. Oral administration of CO-1686 as single agent induces tumor regression in EGFR-mutated NSCLC tumor xenograft and transgenic models. Minimal activity of CO-1686 against the WT EGFR receptor was observed. In NSCLC cells with acquired resistance to CO-1686 in vitro, there was no evidence of additional mutations or amplification of the EGFR gene, but resistant cells exhibited signs of epithelial-mesenchymal transition and demonstrated increased sensitivity to AKT inhibitors. These results suggest that CO-1686 may offer a novel therapeutic option for patients with mutant EGFR NSCLC. SIGNIFICANCE: We report the preclinical development of a novel covalent inhibitor, CO-1686, that irreversibly and selectively inhibits mutant EGFR, in particular the T790M drug-resistance mutation, in NSCLC models. CO-1686 is the fi rst drug of its class in clinical development for the treatment of T790M-positive NSCLC, potentially offering potent inhibition of mutant EGFR while avoiding the on-target toxicity observed with inhibition of the WT EGFR.
Asunto(s)
Acrilamidas/farmacología , Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/genética , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas/farmacología , Acrilamidas/administración & dosificación , Administración Oral , Animales , Antineoplásicos/administración & dosificación , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Ensayos de Selección de Medicamentos Antitumorales , Transición Epitelial-Mesenquimal/efectos de los fármacos , Receptores ErbB/metabolismo , Femenino , Células HEK293 , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Ratones Transgénicos , Terapia Molecular Dirigida , Proteínas Mutantes/antagonistas & inhibidores , Proteínas Mutantes/metabolismo , Inhibidores de Proteínas Quinasas/administración & dosificación , Pirimidinas/administración & dosificación , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
OBJECTIVE: To determine the disease-modifying activity and mechanism of action of the orally available methionine aminopeptidase type 2 inhibitor, [(1R)-1-carbamoyl-2-methyl-propyl]-carbamic acid-(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methyl-but-2-enyl)-oxiranyl]-1-oxa-spiro [2.5] oct-6-yl ester (PPI-2458), in a rat model of peptidoglycan-polysaccharide (PG-PS)-induced arthritis. METHODS: Arthritis was induced in rats by administration of PG-PS, causing tarsal joint swelling and histopathologic changes characteristic of rheumatoid arthritis (RA). PPI-2458, a potent irreversible methionine aminopeptidase type 2 inhibitor, was administered orally every other day at 1, 5, or 10 mg/kg. RESULTS: In an in vitro osteoclastogenesis model, PPI-2458 potently inhibited osteoclast differentiation and bone resorption. In the rat PG-PS arthritis model, PPI-2458 afforded significant protection against established disease after therapeutic dosing. This in vivo activity of PPI-2458 was linked to the inhibition of methionine aminopeptidase type 2. Histopathologic assessment of affected joints showed improvement in processes of inflammation, bone resorption, and cartilage erosion, associated with significant improvement in all clinical indices. The protective effects of PPI-2458 against bone destruction in vivo, including the structural preservation of affected hind joints, correlated with improvements in bone histomorphometric markers, as determined by microfocal computed tomography and a significant decrease in systemic C-telopeptide of type I collagen, suggesting decreased osteoclast activity in vivo. Moreover, PPI-2458 prevented cartilage erosion as shown by a significant decrease in systemic cartilage oligomeric matrix protein. CONCLUSION: The findings of this study suggest that PPI-2458 exerts disease-modifying activity in experimental arthritis through its direct inhibition of several pathophysiologic processes of this disease. These results provide a rationale for assessing the potential of PPI-2458 as a novel RA therapy.