RESUMEN
Thiazolin-4-ones and their derivatives represent important heterocyclic scaffolds with various applications in medicinal chemistry. For that reason, the synthesis of two 5-substituted thiazolidin-4-one derivatives was performed. Their structure assignment was conducted by NMR experiments (2D-COSY, 2D-NOESY, 2D-HSQC and 2D-HMBC) and conformational analysis was conducted through Density Functional Theory calculations and 2D-NOESY. Conformational analysis showed that these two molecules adopt exo conformation. Their global minimum structures have two double bonds (C=N, C=C) in Z conformation and the third double (C=N) in E. Our DFT results are in agreement with the 2D-NMR measurements. Furthermore, the reaction isomerization paths were studied via DFT to check the stability of the conformers. Finally, some potential targets were found through the SwissADME platform and docking experiments were performed. Both compounds bind strongly to five macromolecules (triazoloquinazolines, mglur3, Jak3, Danio rerio HDAC6 CD2, acetylcholinesterase) and via SwissADME it was found that these two molecules obey Lipinski's Rule of Five.
Asunto(s)
Conformación Molecular , Simulación del Acoplamiento Molecular , Tiazolidinas , Tiazolidinas/química , Tiazolidinas/síntesis química , Isomerismo , Animales , Acetilcolinesterasa/química , Acetilcolinesterasa/metabolismo , Pez Cebra , Espectroscopía de Resonancia Magnética , Janus Quinasa 3/antagonistas & inhibidores , Janus Quinasa 3/metabolismo , Janus Quinasa 3/química , Estructura MolecularRESUMEN
JAK3 differs from other JAK family members in terms of tissue distribution and functional properties, making it a promising target for autoimmune disease treatment. However, due to the high homology of these family members, targeting JAK3 selectively is difficult. As a result, exploiting small changes or selectively boosting affinity within the ATP binding region to produce new tailored inhibitors of JAK3 is extremely beneficial. PubChem CID 137321159 was used as the lead inhibitor in this study to preserve the characteristic structure and to collocate it with the redesigned new parent core structure, from which a series of 1,7-dihydro-dipyrrolo [2,3-b:3',2'-e] pyridine derivatives were obtained using the backbone growth method. From the proposed compounds, 14 inhibitors of JAK3 were found based on the docking scoring evaluation. The RMSD and MM/PBSA methods of molecular dynamics simulations were also used to confirm the stable nature of this series of complex systems, and the weak protein−ligand interactions during the dynamics were graphically evaluated and further investigated. The results demonstrated that the new parent core structure fully occupied the hydrophobic cavity, enhanced the interactions of residues LEU828, VAL836, LYS855, GLU903, LEU905 and LEU956, and maintained the structural stability. Apart from this, the results of the analysis show that the binding efficiency of the designed inhibitors of JAK3 is mainly achieved by electrostatic and VDW interactions and the order of the binding free energy with JAK3 is: 8 (−70.286 kJ/mol) > 11 (−64.523 kJ/mol) > 6 (−51.225 kJ/mol) > 17 (−42.822 kJ/mol) > 10 (−40.975 kJ/mol) > 19 (−39.754 kJ/mol). This study may provide a valuable reference for the discovery of novel JAK3 inhibitors for those patients with immune diseases.
Asunto(s)
Enfermedades Autoinmunes , Janus Quinasa 3 , Inhibidores de Proteínas Quinasas , Piridinas , Diseño de Fármacos , Humanos , Janus Quinasa 3/antagonistas & inhibidores , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Simulación de Dinámica Molecular , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/química , Piridinas/farmacologíaRESUMEN
The SH2B family of adaptor proteins, SH2-B, APS, and LNK are key modulators of cellular signalling pathways. Whilst SH2-B and APS have been partially structurally and biochemically characterised, to date there has been no such characterisation of LNK. Here we present two crystal structures of the LNK substrate recognition domain, the SH2 domain, bound to phosphorylated motifs from JAK2 and EPOR, and biochemically define the basis for target recognition. The LNK SH2 domain adopts a canonical SH2 domain fold with an additional N-terminal helix. Targeted analysis of binding to phosphosites in signalling pathways indicated that specificity is conferred by amino acids one- and three-residues downstream of the phosphotyrosine. Several mutations in LNK showed impaired target binding in vitro and a reduced ability to inhibit signalling, allowing an understanding of the molecular basis of LNK dysfunction in variants identified in patients with myeloproliferative disease.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Secuencias de Aminoácidos , Animales , Sitios de Unión , Cristalografía por Rayos X , Humanos , Janus Quinasa 2/química , Janus Quinasa 2/metabolismo , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Ratones , Mutación , Trastornos Mieloproliferativos/genética , Fosfotirosina , Unión Proteica , Proteínas Proto-Oncogénicas c-kit/química , Proteínas Proto-Oncogénicas c-kit/metabolismo , Receptores de Eritropoyetina/química , Receptores de Eritropoyetina/metabolismo , Transducción de Señal , Tirosina Quinasa 3 Similar a fms/química , Tirosina Quinasa 3 Similar a fms/metabolismo , Dominios Homologos srcRESUMEN
Janus Kinases (JAKs) have emerged as an important drug target for the treatment of a number of immune disorders due to the central role that they play in cytokine signalling. 4 isoforms of JAKs exist in mammalian cells and the ideal isoform profile of a JAK inhibitor has been the subject of much debate. JAK3 has been proposed as an ideal target due to its expression being largely restricted to the immune system and its requirement for signalling by cytokine receptors using the common γ-chain. Unlike other JAKs, JAK3 possesses a cysteine in its ATP binding pocket and this has allowed the design of isoform selective covalent JAK3 inhibitors targeting this residue. We report here that mutating this cysteine to serine does not prevent JAK3 catalytic activity but does greatly increase the IC50 for covalent JAK3 inhibitors. Mice with a Cys905Ser knockin mutation in the endogenous JAK3 gene are viable and show no apparent welfare issues. Cells from these mice show normal STAT phosphorylation in response to JAK3 dependent cytokines but are resistant to the effects of covalent JAK3 inhibitors. These mice therefore provide a chemical-genetic model to study JAK3 function.
Asunto(s)
Janus Quinasa 3/genética , Adenosina Trifosfato/metabolismo , Animales , Sitios de Unión , Técnicas de Sustitución del Gen , Humanos , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Ratones , Modelos Genéticos , Dominios Proteicos , Inhibidores de Proteínas Quinasas/químicaRESUMEN
Rheumatoid arthritis (RA) is a systemic autoimmune disorder that commonly affects multiple joints of the body. Currently, there is no permanent cure to the disease, but it can be managed with several potent drugs that cause serious side effects on prolonged use. Traditional remedies are considered promising for the treatment of several diseases, particularly chronic conditions, because they have lower side effects compared to synthetic drugs. In folklore, the rhizome of Alpinia calcarata Roscoe (Zingiberaceae) is used as a major ingredient of herbal formulations to treat RA. Phytoconstituents reported in A. calcarata rhizomes are diterpenoids, sesquiterpenoid, flavonoids, phytosterol, and volatile oils. The present study is intended to understand the molecular-level interaction of phytoconstituents present in A. calcarata rhizomes with RA molecular targets using computational approaches. A total of 30 phytoconstituents reported from the plant were used to carry out docking with 36 known targets of RA. Based on the docking results, 4 flavonoids were found to be strongly interacting with the RA targets. Further, molecular dynamics simulation confirmed stable interaction of quercetin with 6 targets (JAK3, SYK, MMP2, TLR8, IRAK1, and JAK1), galangin with 2 targets (IRAK1 and JAK1), and kaempferol (IRAK1) with one target of RA. Moreover, the presence of these three flavonoids was confirmed in the A. calcarata rhizome extract using LC-MS analysis. The computational study suggests that flavonoids present in A. calcarata rhizome may be responsible for RA modulatory activity. Particularly, quercetin and galangin could be potential development candidates for the treatment of RA. Investigation of Alpinia calcarata constituent interactions with molecular targets of rheumatoid arthritis: docking, molecular dynamics, and network approach.
Asunto(s)
Alpinia/química , Artritis Reumatoide/tratamiento farmacológico , Biología Computacional , Flavonoides/farmacología , Fitoquímicos/farmacología , Artritis Reumatoide/metabolismo , Cromatografía Liquida , Flavonoides/análisis , Flavonoides/química , Humanos , Quinasas Asociadas a Receptores de Interleucina-1/química , Quinasas Asociadas a Receptores de Interleucina-1/efectos de los fármacos , Janus Quinasa 1/química , Janus Quinasa 1/efectos de los fármacos , Janus Quinasa 3/química , Janus Quinasa 3/efectos de los fármacos , Quempferoles/química , Quempferoles/farmacología , Espectrometría de Masas , Metaloproteinasa 2 de la Matriz/química , Metaloproteinasa 2 de la Matriz/efectos de los fármacos , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Fitoquímicos/análisis , Fitoquímicos/química , Extractos Vegetales/química , Quercetina/química , Quercetina/farmacología , Rizoma/química , Quinasa Syk/química , Quinasa Syk/efectos de los fármacos , Receptor Toll-Like 8/química , Receptor Toll-Like 8/efectos de los fármacosRESUMEN
Streptomyces hygroscopicus UFPEDA 3370 was fermented in submerged cultivation and the biomass extract was partitioned, obtaining a fraction purified named EB1. After purification of EB1 fraction, nigericin free acid was obtained and identified. Nigericin presented cytotoxic activity against several cancer cell lines, being most active against HL-60 (human leukemia) and HCT-116 (human colon carcinoma) cell lines, presenting IC50 and (IS) values: 0.0014 µM, (30.0) and 0.0138 µM (3.0), respectively. On HCT-116, nigericin caused apoptosis and autophagy. In this study, nigericin was also screened both in vitro and in silico against a panel of cancer-related kinases. Nigericin was able to inhibit both JAK3 and GSK-3ß kinases in vitro and its binding affinities were mapped through the intermolecular interactions with each target in silico.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias Colorrectales/patología , Nigericina/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Streptomyces/química , Antineoplásicos/química , Antineoplásicos/metabolismo , Apoptosis/efectos de los fármacos , Dominio Catalítico , Línea Celular Tumoral , Humanos , Janus Quinasa 3/antagonistas & inhibidores , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Simulación del Acoplamiento Molecular , Nigericina/química , Nigericina/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/metabolismoRESUMEN
Mutations in the common gamma chain of the interleukin 2 receptor (IL2RG) or the associated downstream signaling enzyme Janus kinase 3 (JAK3) genes are typically characterized by a T cell-negative, B cell-positive, natural killer (NK) cell-negative (T-B+NK-) severe combined immunodeficiency (SCID) immune phenotype. We report clinical course, immunological, genetic and proteomic work-up of two patients with different novel mutations in the IL-2-JAK3 pathway with a rare atypical presentation of T-B+NK- SCID. Lymphocyte subpopulation revealed significant T cells lymphopenia, normal B cells, and NK cells counts (T-B+NK+SCID). Despite the presence of B cells, IgG levels were low and IgA and IgM levels were undetectable. T-cell proliferation in response to mitogens in patient 1 was very low and T-cell receptor V-beta chain repertoire in patient 2 was polyclonal. Whole-exome sequencing revealed novel mutations in both patients (patient 1-c.923delC frame-shift mutation in the IL2RG gene, patient 2-c.G172A a homozygous missense mutation in the JAK3 gene). Bioinformatic analysis of the JAK3 mutation indicated deleterious effect and 3D protein modeling located the mutation to a surface exposed alpha-helix structure. Our findings help to link between genotype and phenotype, which is a key factor for the diagnosis and treatment of SCID patients.
Asunto(s)
Subunidad gamma Común de Receptores de Interleucina/genética , Janus Quinasa 3/genética , Fenotipo , Inmunodeficiencia Combinada Grave/genética , Femenino , Humanos , Lactante , Janus Quinasa 3/química , Masculino , Mutación , Linaje , Conformación Proteica en Hélice alfa , Inmunodeficiencia Combinada Grave/patologíaRESUMEN
Thanks to recent guidelines, the design of safe and effective covalent drugs has gained significant interest. Other than targeting non-conserved nucleophilic residues, optimizing the noncovalent binding framework is important to improve potency and selectivity of covalent binders toward the desired target. Significant efforts have been made in extending the computational toolkits to include a covalent mechanism of protein targeting, like in the development of covalent docking methods for binding mode prediction. To highlight the value of the noncovalent complex in the covalent binding process, here we describe a new protocol using tethered and constrained docking in combination with Dynamic Undocking (DUck) as a tool to privilege strong protein binders for the identification of novel covalent inhibitors. At the end of the protocol, dedicated covalent docking methods were used to rank and select the virtual hits based on the predicted binding mode. By validating the method on JAK3 and KRas, we demonstrate how this fast iterative protocol can be applied to explore a wide chemical space and identify potent targeted covalent inhibitors.
Asunto(s)
Inhibidores Enzimáticos/química , Janus Quinasa 3/química , Proteínas Proto-Oncogénicas p21(ras)/química , Proteínas Recombinantes/química , Bibliotecas de Moléculas Pequeñas/química , Apoptosis , Sitios de Unión , Línea Celular , Supervivencia Celular , Escherichia coli , Humanos , Simulación del Acoplamiento Molecular , Unión Proteica , Conformación Proteica , Programas Informáticos , Relación Estructura-ActividadRESUMEN
Ongoing interest in the discovery of selective JAK3 inhibitors led us to design novel covalent inhibitors that engage the JAK3 residue Cys909 by cyanamide, a structurally and mechanistically differentiated electrophile from other cysteine reacting groups previously incorporated in JAK3 covalent inhibitors. Through crystallography, kinetic, and computational studies, interaction of cyanamide 12 with Cys909 was optimized leading to potent and selective JAK3 inhibitors as exemplified by 32. In relevant cell-based assays and in agreement with previous results from this group, 32 demonstrated that selective inhibition of JAK3 is sufficient to drive JAK1/JAK3-mediated cellular responses. The contribution from extrahepatic processes to the clearance of cyanamide-based covalent inhibitors was also characterized using metabolic and pharmacokinetic data for 12. This work also gave key insights into a productive approach to decrease glutathione/glutathione S-transferase-mediated clearance, a challenge typically encountered during the discovery of covalent kinase inhibitors.
Asunto(s)
Cianamida/química , Cianamida/farmacología , Janus Quinasa 3/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Animales , Cianamida/farmacocinética , Evaluación Preclínica de Medicamentos , Humanos , Concentración 50 Inhibidora , Janus Quinasa 3/química , Masculino , Modelos Moleculares , Conformación Proteica , Inhibidores de Proteínas Quinasas/farmacocinética , Ratas , Distribución TisularRESUMEN
Precursor B-cell acute lymphoblastic leukemia (B-ALL) is associated with recurrent mutations that occur in cancer-initiating cells. There is a need to understand how driver mutations influence clonal evolution of leukemia. The E26-transformation-specific (ETS) transcription factors PU.1 and Spi-B (encoded by Spi1 and Spib) execute a critical role in B-cell development and serve as complementary tumor suppressors. Here, we used a mouse model to conditionally delete Spi1 and Spib genes in developing B cells. These mice developed B-ALL with a median time to euthanasia of 18 weeks. We performed RNA and whole-exome sequencing (WES) on leukemias isolated from Mb1-CreΔPB mice and identified single nucleotide variants (SNVs) in Jak1, Jak3, and Ikzf3 genes, resulting in amino acid sequence changes. Jak3 mutations resulted in amino acid substitutions located in the pseudo-kinase (R653H, V670A) and in the kinase (T844M) domains. Introduction of Jak3 T844M into Spi1/Spib-deficient precursor B cells was sufficient to promote proliferation in response to low IL-7 concentrations in culture, and to promote proliferation and leukemia-like disease in transplanted mice. We conclude that mutations in Janus kinases represent secondary drivers of leukemogenesis that cooperate with Spi1/Spib deletion. This mouse model represents a useful tool to study clonal evolution in B-ALL.
Asunto(s)
Janus Quinasa 1/genética , Janus Quinasa 3/genética , Leucemia Linfocítica Crónica de Células B/patología , Proteínas Proto-Oncogénicas c-ets/genética , Proteínas Proto-Oncogénicas/genética , Transactivadores/genética , Secuencia de Aminoácidos , Animales , Linfocitos B/citología , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Linfocitos B/trasplante , Proliferación Celular , Modelos Animales de Enfermedad , Factor de Transcripción Ikaros , Interleucina-7/farmacología , Janus Quinasa 1/química , Janus Quinasa 3/química , Leucemia Linfocítica Crónica de Células B/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mutagénesis Sitio-Dirigida , Receptores de Interleucina-7/metabolismo , Eliminación de Secuencia , Transactivadores/químicaRESUMEN
Janus kinases are major drivers of immune signaling and have been the focus of anti-inflammatory drug discovery for more than a decade. Because of the invariable colocalization of JAK1 and JAK3 at cytokine receptors, the question if selective JAK3 inhibition is sufficient to effectively block downstream signaling has been highly controversial. Recently, we discovered the covalent-reversible JAK3 inhibitor FM-381 (23) featuring high isoform and kinome selectivity. Crystallography revealed that this inhibitor induces an unprecedented binding pocket by interactions of a nitrile substituent with arginine residues in JAK3. Herein, we describe detailed structure-activity relationships necessary for induction of the arginine pocket and the impact of this structural change on potency, isoform selectivity, and efficacy in cellular models. Furthermore, we evaluated the stability of this novel inhibitor class in in vitro metabolic assays and were able to demonstrate an adequate stability of key compound 23 for in vivo use.
Asunto(s)
Janus Quinasa 3/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Relación Estructura-Actividad , Adenosina Trifosfato/metabolismo , Animales , Sitios de Unión , Células Cultivadas , Cristalografía por Rayos X , Evaluación Preclínica de Medicamentos/métodos , Estabilidad de Medicamentos , Humanos , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Mediciones Luminiscentes/métodos , Ratones , Fosforilación/efectos de los fármacos , Piridinas/química , Factor de Transcripción STAT5/metabolismo , Linfocitos T/efectos de los fármacosRESUMEN
Here, we report the design and synthesis of pyrimidinyl heterocyclic compounds containing terminal electrophiles as irreversible covalent JAK3 inhibitors that exploit a unique cysteine (Cys909) residue in JAK3. Investigation of the structure-activity relationship utilizing kinase assays resulted in the identification of potent and selective JAK3 inhibitors such as T1, T8, T15, T22, and T29. Among them, T29 was verified as a promising JAK3 irreversible inhibitor that possessed the best bioactivity and selectivity against JAKs and kinases containing a cysteine in the residue analogous to Cys909 in JAK3, suggesting that covalent modification of this Cys residue allowed the identification of a highly selective JAK3 inhibitor. Moreover, T29 also displayed a significant anti-inflammatory effect in ICR mice through the inhibition of increased paw thickness, which is worth further optimization to increase its potency and medicinal properties.
Asunto(s)
Antiinflamatorios/síntesis química , Antiinflamatorios/farmacología , Diseño de Fármacos , Janus Quinasa 3/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Animales , Antiinflamatorios/química , Antiinflamatorios/metabolismo , Técnicas de Química Sintética , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Ratones , Ratones Endogámicos ICR , Modelos Moleculares , Dominios Proteicos , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/metabolismo , Relación Estructura-ActividadRESUMEN
Despite increase in the understanding of the pathogenesis of rheumatoid arthritis (RA), it remains a tough challenge. The advent of kinases involved in key intracellular pathways in pathogenesis of RA may provide a new phase of drug discovery for RA. The present study is aimed to identify dual JAK3/[Formula: see text] inhibitors by developing an optimum pharmacophore model integrating the information revealed by ligand-based pharmacophore models and structure-based pharmacophore models (SBPMs). For JAK3 inhibitors, the addition of an aromatic ring feature and for [Formula: see text] the addition of a hydrophobic feature proposed by SBPMs lead to five-point pharmacophore (i.e., AADHR.54 (JAK3)) and six-point pharmacophore (i.e., AAAHRR.45 ([Formula: see text])). The obtained pharmacophores were validated and used for virtual screening and then for docking-based screening. Molecules were further evaluated for ADME properties, and their docked protein complexes were subjected to MM-GBSA energy calculations and molecular dynamic simulations. The top two hit compounds with novel scaffolds 2-oxo-1,2-dihydroquinoline and benzo[d]oxazole showed inhibitory activity for JAK3 and [Formula: see text].
Asunto(s)
Fosfatidilinositol 3-Quinasa Clase I/química , Janus Quinasa 3/química , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Inhibidores de Proteínas Quinasas/química , Sitios de Unión , Fosfatidilinositol 3-Quinasa Clase I/antagonistas & inhibidores , Diseño de Fármacos , Humanos , Janus Quinasa 3/antagonistas & inhibidores , Ligandos , Conformación Molecular , Método de Montecarlo , Unión Proteica , Inhibidores de Proteínas Quinasas/farmacología , Relación Estructura-Actividad Cuantitativa , Reproducibilidad de los ResultadosRESUMEN
The identification of small molecule inhibitors of IRAK4 for the treatment of autoimmune diseases has been an area of intense research. We discovered novel 4,6-diaminonicotinamides which potently inhibit IRAK4. Optimization efforts were aided by X-ray crystal structures of inhibitors bound to IRAK4. Structure activity relationship (SAR) studies led to the identification of compound 29 which exhibited sub-micromolar potency in a LTA stimulated cellular assay.
Asunto(s)
Diseño de Fármacos , Quinasas Asociadas a Receptores de Interleucina-1/antagonistas & inhibidores , Niacinamida/química , Inhibidores de Proteínas Quinasas/química , Sitios de Unión , Cristalografía por Rayos X , Evaluación Preclínica de Medicamentos , Humanos , Concentración 50 Inhibidora , Quinasas Asociadas a Receptores de Interleucina-1/metabolismo , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Conformación Molecular , Simulación de Dinámica Molecular , Niacinamida/metabolismo , Inhibidores de Proteínas Quinasas/metabolismo , Estructura Terciaria de Proteína , Relación Estructura-ActividadRESUMEN
Reversible janus associated kinase (JAK) inhibitors such as tofacitinib and decernotinib block cytokine signaling and are efficacious in treating autoimmune diseases. However, therapeutic doses are limited due to inhibition of other JAK/signal transducer and activator of transcription pathways associated with hematopoiesis, lipid biogenesis, infection, and immune responses. A selective JAK3 inhibitor may have a better therapeutic index; however, until recently, no compounds have been described that maintain JAK3 selectivity in cells, as well as against the kinome, with good physicochemical properties to test the JAK3 hypothesis in vivo. To quantify the biochemical basis for JAK isozyme selectivity, we determined that the apparent Km value for each JAK isozyme ranged from 31.8 to 2.9 µM for JAK1 and JAK3, respectively. To confirm compound activity in cells, we developed a novel enzyme complementation assay that read activity of single JAK isozymes in a cellular context. Reversible JAK3 inhibitors cannot achieve sufficient selectivity against other isozymes in the cellular context due to inherent differences in enzyme ATP Km values. Therefore, we developed irreversible JAK3 compounds that are potent and highly selective in vitro in cells and against the kinome. Compound 2, a potent inhibitor of JAK3 (0.15 nM) was 4300-fold selective for JAK3 over JAK1 in enzyme assays, 67-fold [interleukin (IL)-2 versus IL-6] or 140-fold [IL-2 versus erythropoietin or granulocyte-macrophage colony-stimulating factor (GMCSF)] selective in cellular reporter assays and >35-fold selective in human peripheral blood mononuclear cell assays (IL-7 versus IL-6 or GMCSF). In vivo, selective JAK3 inhibition was sufficient to block the development of inflammation in a rat model of rheumatoid arthritis, while sparing hematopoiesis.
Asunto(s)
Enfermedades Autoinmunes , Janus Quinasa 1 , Janus Quinasa 3 , Piperidinas/farmacología , Pirimidinas/farmacología , Pirroles/farmacología , Animales , Artritis Experimental/tratamiento farmacológico , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/metabolismo , Relación Dosis-Respuesta a Droga , Monitoreo de Drogas/métodos , Humanos , Isoenzimas , Janus Quinasa 1/antagonistas & inhibidores , Janus Quinasa 1/química , Janus Quinasa 1/metabolismo , Janus Quinasa 3/antagonistas & inhibidores , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Monitorización Inmunológica/métodos , Inhibidores de Proteínas Quinasas/farmacología , RatasRESUMEN
To develop more potent JAK3 kinase inhibitors, a series of CP-690550 derivatives were investigated using combined molecular modeling techniques, such as 3D-QSAR, molecular docking and molecular dynamics (MD). The leave-one-out correlation (q2) and non-cross-validated correlation coefficient (r2) of the best CoMFA model are 0.715 and 0.992, respectively. The q2 and r2 values of the best CoMSIA model are 0.739 and 0.995, respectively. The steric, electrostatic, and hydrophobic fields played important roles in determining the inhibitory activity of CP-690550 derivatives. Some new JAK3 kinase inhibitors were designed. Some of them have better inhibitory activity than the most potent Tofacitinib (CP-690550). Molecular docking was used to identify some key amino acid residues at the active site of JAK3 protein. 10ns MD simulations were successfully performed to confirm the detailed binding mode and validate the rationality of docking results. The calculation of the binding free energies by MMPBSA method gives a good correlation with the predicted biological activity. To our knowledge, this is the first report on MD simulations and free energy calculations for this series of compounds. The combination results of this study will be valuable for the development of potent and novel JAK3 kinase inhibitors.
Asunto(s)
Janus Quinasa 3/antagonistas & inhibidores , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Piperidinas/química , Inhibidores de Proteínas Quinasas/química , Pirimidinas/química , Pirroles/química , Relación Estructura-Actividad Cuantitativa , Sitios de Unión , Interacciones Hidrofóbicas e Hidrofílicas , Concentración 50 Inhibidora , Janus Quinasa 3/química , TermodinámicaRESUMEN
The clinical efficacy of multiple kinase inhibitors has caught the interest of Pharmaceutical and Biotech researchers to develop potential drugs with multi-kinase inhibitory activity for complex diseases. In the present work, we attempted to identify dual inhibitors of spleen tyrosine kinase (Syk) and janus kinase 3 (JAK3), keys players in immune signaling, by developing ideal pharmacophores integrating Ligand-based pharmacophore models (LBPMs) and Structure-based pharmacophore models (SBPMs), thereby projecting the optimum pharmacophoric required for inhibition of both the kinases. The four point LBPM; ADPR.14 suggested the presence of one hydrogen bond acceptor, one hydrogen bond donor, one positive ionizable, and one ring aromatic feature for Syk inhibitory activity and AADH.54 proposed the necessity of two hydrogen bond acceptor, one hydrogen bond donor, and one hydrophobic feature for JAK3 inhibitory activity. To our interest, SBPMs identified additional ring aromatic features required for inhibition of both the kinases. For Syk inhibitory activity, the hydrogen bond acceptor feature indicated by LBPM was devoid of forming hydrogen bonding interaction with the hinge region amino acid residue (Ala451). Thus merging the information revealed by both LBPMs and SBPMs, ideal pharmacophore models i.e. ADPRR.14 (Syk) and AADHR.54 (JAK3) were generated. These models after rigorous statistical validation were used for screening of Asinex database. The systematic virtual screening protocol, including pharmacophore and docking-based screening, ADME property, and MM-GBSA energy calculations, retrieved final 10 hits as dual inhibitors of Syk and JAK3. Final 10 hits thus obtained can aid in the development of potential therapeutic agents for autoimmune disorders. Also the top two hits were evaluated against both the enzymes.
Asunto(s)
Janus Quinasa 3/química , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Inhibidores de Proteínas Quinasas/química , Relación Estructura-Actividad Cuantitativa , Quinasa Syk/química , Simulación por Computador , Diseño de Fármacos , Enlace de Hidrógeno , Janus Quinasa 3/antagonistas & inhibidores , Ligandos , Inhibidores de Proteínas Quinasas/farmacología , Reproducibilidad de los Resultados , Quinasa Syk/antagonistas & inhibidoresRESUMEN
Spiroapplanatumines A-Q (1-12, 14-16, 18, and 20), new spiro meroterpenoids respectively bearing a 6/5/7 or 6/5/5 ring system, along with three known compounds, spirolingzhines A, B, and D, were isolated from the fruiting bodies of the fungus Ganoderma applanatum. Their structures including absolute configurations were assigned by using spectroscopic methods, ECD and 13C NMR calculations, and single-crystal X-ray diffraction analysis. Biological evaluation of all the compounds disclosed that compounds 7 and 8 inhibited JAK3 kinase with IC50 values of 7.0 ± 3.2 and 34.8 ± 21.1 µM, respectively.
Asunto(s)
Agaricales/química , Cuerpos Fructíferos de los Hongos/química , Ganoderma/química , Janus Quinasa 3/antagonistas & inhibidores , Terpenos/aislamiento & purificación , Terpenos/farmacología , Triterpenos/aislamiento & purificación , Triterpenos/farmacología , Concentración 50 Inhibidora , Janus Quinasa 3/química , Espectroscopía de Resonancia Magnética , Estructura Molecular , Terpenos/química , Triterpenos/química , Difracción de Rayos XRESUMEN
Janus kinases (JAKs) are a family of cytoplasmatic tyrosine kinases that are attractive targets for the development of anti-inflammatory drugs given their roles in cytokine signaling. One question regarding JAKs and their inhibitors that remains under intensive debate is whether JAK inhibitors should be isoform selective. Since JAK3 functions are restricted to immune cells, an isoform-selective inhibitor for JAK3 could be especially valuable to achieve clinically more useful and precise effects. However, the high degree of structural conservation makes isoform-selective targeting a challenging task. Here, we present picomolar inhibitors with unprecedented kinome-wide selectivity for JAK3. Selectivity was achieved by concurrent covalent reversible targeting of a JAK3-specific cysteine residue and a ligand-induced binding pocket. We confirmed that in vitro activity and selectivity translate well into the cellular environment and suggest that our inhibitors are powerful tools to elucidate JAK3-specific functions.
Asunto(s)
Janus Quinasa 3/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Sitios de Unión/efectos de los fármacos , Descubrimiento de Drogas , Humanos , Janus Quinasa 3/química , Janus Quinasa 3/metabolismo , Simulación del Acoplamiento Molecular , Transducción de Señal/efectos de los fármacos , Linfocitos T/efectos de los fármacos , Linfocitos T/metabolismoRESUMEN
While several therapeutic options exist, the need for more effective, safe, and convenient treatment for a variety of autoimmune diseases persists. Targeting the Janus tyrosine kinases (JAKs), which play essential roles in cell signaling responses and can contribute to aberrant immune function associated with disease, has emerged as a novel and attractive approach for the development of new autoimmune disease therapies. We screened our compound library against JAK3, a key signaling kinase in immune cells, and identified multiple scaffolds showing good inhibitory activity for this kinase. A particular scaffold of interest, the 1H-pyrrolo[2,3-b]pyridine series (7-azaindoles), was selected for further optimization in part on the basis of binding affinity (Ki) as well as on the basis of cellular potency. Optimization of this chemical series led to the identification of VX-509 (decernotinib), a novel, potent, and selective JAK3 inhibitor, which demonstrates good efficacy in vivo in the rat host versus graft model (HvG). On the basis of these findings, it appears that VX-509 offers potential for the treatment of a variety of autoimmune diseases.