RESUMEN
Cyclic dinucleotides (CDN) and Toll-like receptor (TLR) ligands mobilize antitumor responses by natural killer (NK) cells and T cells, potentially serving as complementary therapies to immune checkpoint therapy. In the clinic thus far, however, CDN therapy targeting stimulator of interferon genes (STING) protein has yielded mixed results, perhaps because it initiates responses potently but does not provide signals to sustain activation and proliferation of activated cytotoxic lymphocytes. To improve efficacy, we combined CDN with a half life-extended interleukin-2 (IL-2) superkine, H9-MSA (mouse serum albumin). CDN/H9-MSA therapy induced dramatic long-term remissions of the most difficult to treat major histocompatibility complex class I (MHC I)deficient and MHC I+ tumor transplant models. H9-MSA combined with CpG oligonucleotide also induced potent responses. Mechanistically, tumor elimination required CD8 T cells and not NK cells in the case of MHC I+ tumors and NK cells but not CD8 T cells in the case of MHC-deficient tumors. Furthermore, combination therapy resulted in more prolonged and more intense NK cell activation, cytotoxicity, and expression of cytotoxic effector molecules in comparison with monotherapy. Remarkably, in a primary autochthonous sarcoma model that is refractory to PD-1 checkpoint therapy, the combination of CDN/H9-MSA with checkpoint therapy yielded long-term remissions in the majority of the animals, mediated by T cells and NK cells. This combination therapy has the potential to activate responses in tumors resistant to current therapies and prevent MHC I loss accompanying acquired resistance of tumors to checkpoint therapy.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica , Antígenos de Histocompatibilidad Clase I , Inmunoterapia , Interleucina-2 , Proteínas de la Membrana , Neoplasias , Nucleótidos Cíclicos , Oligodesoxirribonucleótidos , Albúmina Sérica , Animales , Linfocitos T CD8-positivos/inmunología , Antígenos de Histocompatibilidad Clase I/genética , Humanos , Inmunoterapia/métodos , Interleucina-2/inmunología , Células Asesinas Naturales/inmunología , Proteínas de la Membrana/agonistas , Ratones , Neoplasias/genética , Neoplasias/terapia , Nucleótidos Cíclicos/uso terapéutico , Oligodesoxirribonucleótidos/uso terapéutico , Albúmina Sérica/uso terapéuticoRESUMEN
The ubiquitin system regulates essential cellular processes in eukaryotes. Ubiquitin is ligated to substrate proteins as monomers or chains and the topology of ubiquitin modifications regulates substrate interactions with specific proteins. Thus ubiquitination directs a variety of substrate fates including proteasomal degradation. Deubiquitinase enzymes cleave ubiquitin from substrates and are implicated in disease; for example, ubiquitin-specific protease-7 (USP7) regulates stability of the p53 tumour suppressor and other proteins critical for tumour cell survival. However, developing selective deubiquitinase inhibitors has been challenging and no co-crystal structures have been solved with small-molecule inhibitors. Here, using nuclear magnetic resonance-based screening and structure-based design, we describe the development of selective USP7 inhibitors GNE-6640 and GNE-6776. These compounds induce tumour cell death and enhance cytotoxicity with chemotherapeutic agents and targeted compounds, including PIM kinase inhibitors. Structural studies reveal that GNE-6640 and GNE-6776 non-covalently target USP7 12 Å distant from the catalytic cysteine. The compounds attenuate ubiquitin binding and thus inhibit USP7 deubiquitinase activity. GNE-6640 and GNE-6776 interact with acidic residues that mediate hydrogen-bond interactions with the ubiquitin Lys48 side chain, suggesting that USP7 preferentially interacts with and cleaves ubiquitin moieties that have free Lys48 side chains. We investigated this idea by engineering di-ubiquitin chains containing differential proximal and distal isotopic labels and measuring USP7 binding by nuclear magnetic resonance. This preferential binding protracted the depolymerization kinetics of Lys48-linked ubiquitin chains relative to Lys63-linked chains. In summary, engineering compounds that inhibit USP7 activity by attenuating ubiquitin binding suggests opportunities for developing other deubiquitinase inhibitors and may be a strategy more broadly applicable to inhibiting proteins that require ubiquitin binding for full functional activity.
Asunto(s)
Aminopiridinas/química , Aminopiridinas/farmacología , Indazoles/química , Indazoles/farmacología , Fenoles/química , Fenoles/farmacología , Piridinas/química , Piridinas/farmacología , Peptidasa Específica de Ubiquitina 7/antagonistas & inhibidores , Ubiquitina/metabolismo , Animales , Unión Competitiva , Línea Celular Tumoral , Sinergismo Farmacológico , Femenino , Humanos , Ratones , Ratones SCID , Modelos Moleculares , Neoplasias/tratamiento farmacológico , Neoplasias/enzimología , Neoplasias/patología , Unión Proteica , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Especificidad por Sustrato , Proteína p53 Supresora de Tumor/deficiencia , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Ubiquitina/química , Peptidasa Específica de Ubiquitina 7/química , Peptidasa Específica de Ubiquitina 7/deficiencia , Peptidasa Específica de Ubiquitina 7/metabolismoRESUMEN
Cell migration is a stepwise process that coordinates multiple molecular machineries. Using in vitro angiogenesis screens with short interfering RNA and chemical inhibitors, we define here a MAP4K4-moesin-talin-ß1-integrin molecular pathway that promotes efficient plasma membrane retraction during endothelial cell migration. Loss of MAP4K4 decreased membrane dynamics, slowed endothelial cell migration, and impaired angiogenesis in vitro and in vivo. In migrating endothelial cells, MAP4K4 phosphorylates moesin in retracting membranes at sites of focal adhesion disassembly. Epistasis analyses indicated that moesin functions downstream of MAP4K4 to inactivate integrin by competing with talin for binding to ß1-integrin intracellular domain. Consequently, loss of moesin (encoded by the MSN gene) or MAP4K4 reduced adhesion disassembly rate in endothelial cells. Additionally, α5ß1-integrin blockade reversed the membrane retraction defects associated with loss of Map4k4 in vitro and in vivo. Our study uncovers a novel aspect of endothelial cell migration. Finally, loss of MAP4K4 function suppressed pathological angiogenesis in disease models, identifying MAP4K4 as a potential therapeutic target.
Asunto(s)
Movimiento Celular , Células Endoteliales/citología , Células Endoteliales/metabolismo , Integrinas/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Secuencias de Aminoácidos , Animales , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Forma de la Célula/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Epistasis Genética , Adhesiones Focales/metabolismo , Humanos , Integrina alfa1/efectos de los fármacos , Integrina alfa1/metabolismo , Integrina beta1/química , Integrina beta1/efectos de los fármacos , Integrina beta1/metabolismo , Integrinas/efectos de los fármacos , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/deficiencia , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Ratones , Proteínas de Microfilamentos/deficiencia , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Neovascularización Patológica , Fosforilación , Unión Proteica , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Estructura Terciaria de Proteína , Talina/química , Talina/metabolismoRESUMEN
MAP4K4 has been shown to regulate key cellular processes that are tied to disease pathogenesis. In an effort to generate small molecule MAP4K4 inhibitors, a fragment-based screen was carried out and a pyrrolotriazine fragment with excellent ligand efficiency was identified. Further modification of this fragment guided by X-ray crystal structures and molecular modeling led to the discovery of a series of promising compounds with good structural diversity and physicochemical properties. These compounds exhibited single digit nanomolar potency and compounds 35 and 44 achieved good in vivo exposure.
Asunto(s)
Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Triazinas/farmacología , Animales , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ligandos , Ratones , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Relación Estructura-Actividad , Triazinas/síntesis química , Triazinas/química , Quinasa de Factor Nuclear kappa BRESUMEN
A series of suitable five-membered heterocyclic alternatives to thiophenes within a thienobenzoxepin class of PI3-kinase (PI3K) inhibitors was discovered. Specific thiazolobenzoxepin 8-substitution was identified that increased selectivity over PI3Kß. PI3Kß-sparing compound 27 (PI3Kß Ki,app/PI3Kα Ki,app=57) demonstrated dose-dependent knockdown of pAKT, pPRAS40 and pS6RP in vivo as well as differential effects in an in vitro proliferation cell line screen compared to pan PI3K inhibitor GDC-0941. A new structure-based hypothesis for reducing inhibition of the PI3K ß isoform while maintaining activity against α, δ and γ isoforms is presented.
Asunto(s)
Benzoxepinas/química , Inhibidores Enzimáticos/química , Inhibidores de las Quinasa Fosfoinosítidos-3 , Tiazoles/química , Benzoxepinas/síntesis química , Benzoxepinas/farmacología , Sitios de Unión , Proliferación Celular/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Humanos , Células MCF-7 , Simulación del Acoplamiento Molecular , Fosfatidilinositol 3-Quinasa/metabolismo , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/metabolismo , Estructura Terciaria de Proteína , Proteínas Proto-Oncogénicas c-akt/metabolismo , Relación Estructura-ActividadRESUMEN
Several immunotherapy approaches that mobilize CD8+ T cell responses stimulate tumor rejection, and some, such as checkpoint blockade, have been approved for several cancer indications and show impressive increases in patient survival. However, tumors may evade CD8+ T cell recognition via loss of MHC molecules or because they contain few or no neoantigens. Therefore, approaches are needed to combat CD8+ T cell-resistant cancers. STING-activating cyclic dinucleotides (CDNs) are a new class of immune-stimulating agents that elicit impressive CD8+ T cell-mediated tumor rejection in preclinical tumor models and are now being tested in clinical trials. Here, we demonstrate powerful CDN-induced, natural killer (NK) cell-mediated tumor rejection in numerous tumor models, independent of CD8+ T cells. CDNs enhanced NK cell activation, cytotoxicity, and antitumor effects in part by inducing type I interferon (IFN). IFN acted in part directly on NK cells in vivo and in part indirectly via the induction of IL-15 and IL-15 receptors, which were important for CDN-induced NK activation and tumor control. After in vivo administration of CDNs, dendritic cells (DCs) up-regulated IL-15Rα in an IFN-dependent manner. Mice lacking the type I IFN receptor specifically on DCs had reduced NK cell activation and tumor control. Therapeutics that activate NK cells, such as CDNs, checkpoint inhibitors, NK cell engagers, and cytokines, may represent next-generation approaches to cancer immunotherapy.
Asunto(s)
Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Células Asesinas Naturales/inmunología , Proteínas de la Membrana/agonistas , Neoplasias/tratamiento farmacológico , Neoplasias/inmunología , Animales , Sistemas CRISPR-Cas/efectos de los fármacos , Sistemas CRISPR-Cas/inmunología , Interferón Tipo I/farmacología , Células Asesinas Naturales/efectos de los fármacos , Proteínas de la Membrana/inmunología , Ratones , Ratones Congénicos , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Neoplasias/patología , Células Tumorales CultivadasRESUMEN
The adenosinergic pathway represents an attractive new therapeutic approach in cancer immunotherapy. In this pathway, ecto-5-nucleotidase CD73 has the unique function of regulating production of immunosuppressive adenosine (ADO) through the hydrolysis of AMP. CD73 is overexpressed in many cancers, resulting in elevated levels of ADO that correspond to poor patient prognosis. Therefore, reducing the level of ADO via inhibition of CD73 is a potential strategy for treating cancers. Based on the binding mode of adenosine 5'-(α,ß-methylene)diphosphate (AOPCP) with human CD73, we designed a series of novel monophosphonate small-molecule CD73 inhibitors. Among them, OP-5244 (35) proved to be a highly potent and orally bioavailable CD73 inhibitor. In preclinical studies, 35 completely inhibited ADO production in both human cancer cells and CD8+ T cells. Furthermore, 35 lowered the ratio of ADO/AMP significantly and reversed immunosuppression in mouse models, indicating its potential as an in vivo tool compound for further development.
Asunto(s)
5'-Nucleotidasa/antagonistas & inhibidores , Adenosina/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Factores Inmunológicos/farmacología , Nucleósidos/farmacología , Organofosfonatos/farmacología , Administración Oral , Animales , Linfocitos T CD8-positivos/efectos de los fármacos , Línea Celular Tumoral , Perros , Inhibidores Enzimáticos/administración & dosificación , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacocinética , Proteínas Ligadas a GPI/antagonistas & inhibidores , Humanos , Factores Inmunológicos/administración & dosificación , Factores Inmunológicos/síntesis química , Factores Inmunológicos/farmacocinética , Macaca fascicularis , Ratones Endogámicos BALB C , Estructura Molecular , Nucleósidos/administración & dosificación , Nucleósidos/síntesis química , Nucleósidos/farmacocinética , Organofosfonatos/administración & dosificación , Organofosfonatos/síntesis química , Organofosfonatos/farmacocinética , Ratas , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
The xishacorene natural products are structurally unique apolar diterpenoids that feature a bicyclo[3.3.1] framework. These secondary metabolites likely arise from the well-studied, structurally related diterpenoid fuscol. In this manuscript, we describe the conversion of fuscol to xishacorenes A, B, and C, as well as a previously unreported congener, which we have named xishacorene D. In addition, we describe immunomodulatory activity studies of the xishacorenes, a structurally related analogue, and fuscol. These studies were aided by an accurate determination of the physical properties (e.g., molar extinction coefficient) of the highly apolar xishacorenes.
RESUMEN
A series of pyrazolopyrimidine inhibitors of IRAK4 were developed from a high-throughput screen (HTS). Modification of an HTS hit led to a series of bicyclic heterocycles with improved potency and kinase selectivity but lacking sufficient solubility to progress in vivo. Structure-based drug design, informed by cocrystal structures with the protein and small-molecule crystal structures, yielded a series of dihydrobenzofurans. This semisaturated bicycle provided superior druglike properties while maintaining excellent potency and selectivity. Improved physicochemical properties allowed for progression into in vivo experiments, where lead molecules exhibited low clearance and showed target-based inhibition of IRAK4 signaling in an inflammation-mediated PK/PD mouse model.
Asunto(s)
Quinasas Asociadas a Receptores de Interleucina-1/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles/farmacología , Pirimidinas/farmacología , Aminoquinolinas/síntesis química , Aminoquinolinas/metabolismo , Aminoquinolinas/farmacología , Animales , Benzofuranos/síntesis química , Benzofuranos/metabolismo , Benzofuranos/farmacología , Dominio Catalítico , Femenino , Humanos , Quinasas Asociadas a Receptores de Interleucina-1/metabolismo , Ratones Endogámicos C57BL , Estructura Molecular , Unión Proteica , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/metabolismo , Pirazoles/síntesis química , Pirazoles/metabolismo , Pirimidinas/síntesis química , Pirimidinas/metabolismo , Ratas , Transducción de Señal/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
Tumor neoantigens arising from somatic mutations in the cancer genome are less likely to be subject to central immune tolerance and are therefore attractive targets for vaccine immunotherapy. We utilized whole-exome sequencing, RNA sequencing (RNASeq), and an in silico immunogenicity prediction algorithm, NetMHC, to generate a neoantigen-targeted vaccine, PancVAX, which was administered together with the STING adjuvant ADU-V16 to mice bearing pancreatic adenocarcinoma (Panc02) cells. PancVAX activated a neoepitope-specific T cell repertoire within the tumor and caused transient tumor regression. When given in combination with two checkpoint modulators, namely anti-PD-1 and agonist OX40 antibodies, PancVAX resulted in enhanced and more durable tumor regression and a survival benefit. The addition of OX40 to vaccine reduced the coexpression of T cell exhaustion markers, Lag3 and PD-1, and resulted in rejection of tumors upon contralateral rechallenge, suggesting the induction of T cell memory. Together, these data provide the framework for testing personalized neoantigen-based combinatorial vaccine strategies in patients with pancreatic and other nonimmunogenic cancers.
Asunto(s)
Adenocarcinoma/terapia , Antineoplásicos Inmunológicos/farmacología , Vacunas contra el Cáncer/administración & dosificación , Inmunoterapia/métodos , Neoplasias Pancreáticas/terapia , Adenocarcinoma/genética , Adenocarcinoma/inmunología , Adyuvantes Inmunológicos/administración & dosificación , Animales , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/inmunología , Antineoplásicos Inmunológicos/uso terapéutico , Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Línea Celular Tumoral/trasplante , Terapia Combinada/métodos , Modelos Animales de Enfermedad , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Humanos , Inmunogenicidad Vacunal , Proteínas de la Membrana/inmunología , Ratones , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/inmunología , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Receptor de Muerte Celular Programada 1/inmunología , Receptores OX40/agonistas , Receptores OX40/inmunología , Resultado del Tratamiento , Escape del Tumor/efectos de los fármacos , Escape del Tumor/inmunología , Vacunas de Subunidad/administración & dosificación , Vacunas de Subunidad/genética , Vacunas de Subunidad/inmunologíaRESUMEN
Intratumoral (IT) STING activation results in tumor regression in preclinical models, yet factors dictating the balance between innate and adaptive anti-tumor immunity are unclear. Here, clinical candidate STING agonist ADU-S100 (S100) is used in an IT dosing regimen optimized for adaptive immunity to uncover requirements for a T cell-driven response compatible with checkpoint inhibitors (CPIs). In contrast to high-dose tumor ablative regimens that result in systemic S100 distribution, low-dose immunogenic regimens induce local activation of tumor-specific CD8+ effector T cells that are responsible for durable anti-tumor immunity and can be enhanced with CPIs. Both hematopoietic cell STING expression and signaling through IFNAR are required for tumor-specific T cell activation, and in the context of optimized T cell responses, TNFα is dispensable for tumor control. In a poorly immunogenic model, S100 combined with CPIs generates a survival benefit and durable protection. These results provide fundamental mechanistic insights into STING-induced anti-tumor immunity.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Inmunidad , Proteínas de la Membrana/metabolismo , Neoplasias/inmunología , Animales , Antígeno CTLA-4/metabolismo , Línea Celular Tumoral , Citocinas/metabolismo , Relación Dosis-Respuesta Inmunológica , Resistencia a Antineoplásicos , Hematopoyesis , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Neoplasias/patología , Receptor de Muerte Celular Programada 1/metabolismo , Proteínas S100/administración & dosificación , Proteínas S100/inmunologíaRESUMEN
USP7 is a deubiquitinase implicated in destabilizing the tumor suppressor p53, and for this reason it has gained increasing attention as a potential oncology target for small molecule inhibitors. Herein we describe the biophysical, biochemical, and computational approaches that led to the identification of 4-(2-aminopyridin-3-yl)phenol compounds described by Kategaya ( Nature 2017 , 550 , 534 - 538 ) as specific inhibitors of USP7. Fragment based lead discovery (FBLD) by NMR combined with virtual screening and re-mining of biochemical high-throughput screening (HTS) hits led to the discovery of a series of ligands that bind in the "palm" region of the catalytic domain of USP7 and inhibit its catalytic activity. These ligands were then optimized by structure-based design to yield cell-active molecules with reasonable physical properties. This discovery process not only involved multiple techniques working in concert but also illustrated a unique way in which hits from orthogonal screening approaches complemented each other for lead identification.
Asunto(s)
Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Bibliotecas de Moléculas Pequeñas/farmacología , Peptidasa Específica de Ubiquitina 7/antagonistas & inhibidores , Aminopiridinas/química , Sitios de Unión , Dominio Catalítico , Línea Celular , Simulación por Computador , Cristalografía por Rayos X , Descubrimiento de Drogas/métodos , Evaluación Preclínica de Medicamentos/métodos , Humanos , Espectroscopía de Resonancia Magnética/métodos , Oxadiazoles/química , Bibliotecas de Moléculas Pequeñas/síntesis química , Bibliotecas de Moléculas Pequeñas/química , Peptidasa Específica de Ubiquitina 7/química , Peptidasa Específica de Ubiquitina 7/metabolismoRESUMEN
Inhibition of phosphoinositide 3-kinase (PI3K) signaling is an appealing approach to treat brain tumors, especially glioblastoma multiforme (GBM). We previously disclosed our successful approach to prospectively design potent and blood-brain barrier (BBB) penetrating PI3K inhibitors. The previously disclosed molecules were ultimately deemed not suitable for clinical development due to projected poor metabolic stability in humans. We, therefore, extended our studies to identify a BBB penetrating inhibitor of PI3K that was also projected to be metabolically stable in human. These efforts required identification of a distinct scaffold for PI3K inhibitors relative to our previous efforts and ultimately resulted in the identification of GDC-0084 (16). The discovery and preclinical characterization of this molecule are described within.
RESUMEN
Inhibitors of the class I phosphoinositide 3-kinase (PI3K) isoform PI3Kα have received substantial attention for their potential use in cancer therapy. Despite the particular attraction of targeting PI3Kα, achieving selectivity for the inhibition of this isoform has proved challenging. Herein we report the discovery of inhibitors of PI3Kα that have selectivity over the other class I isoforms and all other kinases tested. In GDC-0032 (3, taselisib), we previously minimized inhibition of PI3Kß relative to the other class I insoforms. Subsequently, we extended our efforts to identify PI3Kα-specific inhibitors using PI3Kα crystal structures to inform the design of benzoxazepin inhibitors with selectivity for PI3Kα through interactions with a nonconserved residue. Several molecules selective for PI3Kα relative to the other class I isoforms, as well as other kinases, were identified. Optimization of properties related to drug metabolism then culminated in the identification of the clinical candidate GDC-0326 (4).
Asunto(s)
Antineoplásicos/farmacología , Benzoxepinas/farmacología , Diseño de Fármacos , Imidazoles/farmacología , Oxazepinas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/metabolismo , Benzoxepinas/química , Benzoxepinas/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Fosfatidilinositol 3-Quinasa Clase I , Perros , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Imidazoles/química , Imidazoles/metabolismo , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Macaca fascicularis , Ratones , Microsomas Hepáticos/química , Microsomas Hepáticos/metabolismo , Modelos Moleculares , Estructura Molecular , Oxazepinas/química , Oxazepinas/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/metabolismo , Ratas , Ratas Sprague-Dawley , Relación Estructura-ActividadRESUMEN
p21-activated kinase 1 (PAK1) has an important role in transducing signals in several oncogenic pathways. The concept of inhibiting this kinase has garnered significant interest over the past decade, particularly for targeting cancers associated with PAK1 amplification. Animal studies with the selective group I PAK (pan-PAK1, 2, 3) inhibitor G-5555 from the pyrido[2,3-d]pyrimidin-7-one class uncovered acute toxicity with a narrow therapeutic window. To attempt mitigating the toxicity, we introduced significant structural changes, culminating in the discovery of the potent pyridone side chain analogue G-9791. Mouse tolerability studies with this compound, other members of this series, and compounds from two structurally distinct classes revealed persistent toxicity and a correlation of minimum toxic concentrations and PAK1/2 mediated cellular potencies. Broad screening of selected PAK inhibitors revealed PAK1, 2, and 3 as the only overlapping targets. Our data suggest acute cardiovascular toxicity resulting from the inhibition of PAK2, which may be enhanced by PAK1 inhibition, and cautions against continued pursuit of pan-group I PAK inhibitors in drug discovery.
Asunto(s)
Enfermedades Cardiovasculares/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/farmacología , Pirimidinas/farmacología , Quinasas p21 Activadas/antagonistas & inhibidores , Enfermedad Aguda , Animales , Relación Dosis-Respuesta a Droga , Femenino , Ratones , Ratones Endogámicos BALB C , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Piridinas/síntesis química , Piridinas/química , Piridonas , Pirimidinas/síntesis química , Pirimidinas/química , Relación Estructura-Actividad , Quinasas p21 Activadas/metabolismoRESUMEN
[reaction: see text] Ni-catalyzed reductive coupling of aryl alkynes (1) and enantiomerically enriched alpha-oxyaldehydes (2) afford differentiated anti-1,2-diols (3) with high diastereoselectivity and regioselectivity, despite the fact that the methoxymethyl (MOM) and para-methoxybenzyl (PMB) protective groups typically favor syn-1,2-diol formation in carbonyl addition reactions of this family of aldehydes.
Asunto(s)
Alcoholes/síntesis química , Aldehídos/química , Alquinos/química , Níquel/química , Alcoholes/química , Catálisis , Estructura Molecular , Oxidación-Reducción , EstereoisomerismoRESUMEN
The diverse roles of deubiquitinating enzymes, or DUBs, in determining the fate of specific proteins continue to unfold. Concurrent with the revelation of DUBs as potential therapeutic targets are publications of small molecule inhibitors of these enzymes. In this review, we summarize these molecules and their associated data and suggest additional experiments to further validate and characterize these compounds. We believe the field of drug discovery against DUBs is still in its infancy, but advances in assay development, biophysical techniques, and screening libraries hold promise for identifying suitable agents that could advance into the clinic.
Asunto(s)
Inhibidores Enzimáticos/farmacología , Proteasas Ubiquitina-Específicas/antagonistas & inhibidores , Cristalografía por Rayos X , Descubrimiento de Drogas , Inhibidores Enzimáticos/química , Humanos , Modelos Moleculares , Estructura Molecular , Relación Estructura-Actividad , Proteasas Ubiquitina-Específicas/metabolismoRESUMEN
Diverse biological roles for mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) have necessitated the identification of potent inhibitors in order to study its function in various disease contexts. In particular, compounds that can be used to carry out such studies in vivo would be critical for elucidating the potential for therapeutic intervention. A structure-based design effort coupled with property-guided optimization directed at minimizing the ability of the inhibitors to cross into the CNS led to an advanced compound 13 (GNE-495) that showed excellent potency and good PK and was used to demonstrate in vivo efficacy in a retinal angiogenesis model recapitulating effects that were observed in the inducible Map4k4 knockout mice.
RESUMEN
Signaling pathways intersecting with the p21-activated kinases (PAKs) play important roles in tumorigenesis and cancer progression. By recognizing that the limitations of FRAX1036 (1) were chiefly associated with the highly basic amine it contained, we devised a mitigation strategy to address several issues such as hERG activity. The 5-amino-1,3-dioxanyl moiety was identified as an effective means of reducing pK a and logP simultaneously. When positioned properly within the scaffold, this group conferred several benefits including potency, pharmacokinetics, and selectivity. Mouse xenograft PK/PD studies were carried out using an advanced compound, G-5555 (12), derived from this approach. These studies concluded that dose-dependent pathway modulation was achievable and paves the way for further in vivo investigations of PAK1 function in cancer and other diseases.
RESUMEN
Mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) is a serine/threonine kinase implicated in the regulation of many biological processes. A fragment-based lead discovery approach was used to generate potent and selective MAP4K4 inhibitors. The fragment hit pursued in this article had excellent ligand efficiency (LE), an important attribute for subsequent successful optimization into drug-like lead compounds. The optimization efforts eventually led us to focus on the pyridopyrimidine series, from which 6-(2-fluoropyridin-4-yl)pyrido[3,2-d]pyrimidin-4-amine (29) was identified. This compound had low nanomolar potency, excellent kinase selectivity, and good in vivo exposure, and demonstrated in vivo pharmacodynamic effects in a human tumor xenograft model.