RESUMO
Several series of thieno[2-3-b]pyridine analogues were synthesized and screened for inhibitory activity against eukaryotic elongation factor-2 kinase (eEF2-K). Modifications around several regions of the lead molecules were made, with a ring fusion adjacent to the nitrogen on the thienopyridine core being critical for activity. The most active compound 34 shows an IC(50) of 170 nM against eEF2-K in vitro.
Assuntos
Quinase do Fator 2 de Elongação/antagonistas & inibidores , Quinase do Fator 2 de Elongação/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Piridinas/química , Piridinas/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Relação Estrutura-AtividadeRESUMO
Valosin-containing protein (VCP; also known as p97) is a member of the AAA ATPase family with a central role in the ubiquitin-degradation of misfolded proteins. VCP also exhibits antiapoptotic function and metastasis via activation of nuclear factor kappa-B signaling pathway. We have discovered that 2-anilino-4-aryl-1,3-thiazoles are potent drug-like inhibitors of this enzyme. The identified compounds show low nanomolar VCP potency, demonstrate SAR trends, and show activity in a mechanism based cellular assay. This series of compounds represents the first steps towards a novel, small molecule VCP inhibitor as a cancer therapeutic.
Assuntos
Adenosina Trifosfatases/antagonistas & inibidores , Compostos de Anilina/química , Antineoplásicos/química , Proteínas de Ciclo Celular/antagonistas & inibidores , Tiazóis/química , Adenosina Trifosfatases/metabolismo , Compostos de Anilina/síntese química , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Proteínas de Ciclo Celular/metabolismo , Células HeLa , Humanos , NF-kappa B/metabolismo , Transdução de Sinais , Relação Estrutura-Atividade , Tiazóis/síntese química , Tiazóis/farmacologia , Proteína com ValosinaRESUMO
The devastating sequelae of diabetes mellitus include microvascular permeability, which results in retinopathy. Despite clinical and scientific advances, there remains a need for new approaches to treat retinopathy. Here, we have presented a possible treatment strategy, whereby targeting the small GTPase ARF6 alters VEGFR2 trafficking and reverses signs of pathology in 4 animal models that represent features of diabetic retinopathy and in a fifth model of ocular pathological angiogenesis. Specifically, we determined that the same signaling pathway utilizes distinct GEFs to sequentially activate ARF6, and these GEFs exert distinct but complementary effects on VEGFR2 trafficking and signal transduction. ARF6 activation was independently regulated by 2 different ARF GEFs - ARNO and GEP100. Interaction between VEGFR2 and ARNO activated ARF6 and stimulated VEGFR2 internalization, whereas a VEGFR2 interaction with GEP100 activated ARF6 to promote VEGFR2 recycling via coreceptor binding. Intervening in either pathway inhibited VEGFR2 signal output. Finally, using a combination of in vitro, cellular, genetic, and pharmacologic techniques, we demonstrated that ARF6 is pivotal in VEGFR2 trafficking and that targeting ARF6-mediated VEGFR2 trafficking has potential as a therapeutic approach for retinal vascular diseases such as diabetic retinopathy.
Assuntos
Fatores de Ribosilação do ADP/metabolismo , Retinopatia Diabética/metabolismo , Transdução de Sinais , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Fator 6 de Ribosilação do ADP , Fatores de Ribosilação do ADP/genética , Linhagem Celular , Retinopatia Diabética/genética , Retinopatia Diabética/patologia , Proteínas Ativadoras de GTPase/genética , Proteínas Ativadoras de GTPase/metabolismo , Humanos , Transporte Proteico , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genéticaRESUMO
Activating mutations in Gαq proteins, which form the α subunit of certain heterotrimeric G proteins, drive uveal melanoma oncogenesis by triggering multiple downstream signaling pathways, including PLC/PKC, Rho/Rac, and YAP. Here we show that the small GTPase ARF6 acts as a proximal node of oncogenic Gαq signaling to induce all of these downstream pathways as well as ß-catenin signaling. ARF6 activates these diverse pathways through a common mechanism: the trafficking of GNAQ and ß-catenin from the plasma membrane to cytoplasmic vesicles and the nucleus, respectively. Blocking ARF6 with a small-molecule inhibitor reduces uveal melanoma cell proliferation and tumorigenesis in a mouse model, confirming the functional relevance of this pathway and suggesting a therapeutic strategy for Gα-mediated diseases.
Assuntos
Fatores de Ribosilação do ADP/metabolismo , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/genética , Melanoma/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/administração & dosagem , Neoplasias Uveais/tratamento farmacológico , beta Catenina/metabolismo , Fator 6 de Ribosilação do ADP , Fatores de Ribosilação do ADP/antagonistas & inibidores , Fatores de Ribosilação do ADP/genética , Animais , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Citoplasma/metabolismo , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/metabolismo , Humanos , Melanoma/genética , Melanoma/metabolismo , Camundongos , Transplante de Neoplasias , Transporte Proteico/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Neoplasias Uveais/genética , Neoplasias Uveais/metabolismoRESUMO
ß-Catenin has a dual function in cells: fortifying cadherin-based adhesion at the plasma membrane and activating transcription in the nucleus. We found that in melanoma cells, WNT5A stimulated the disruption of N-cadherin and ß-catenin complexes by activating the guanosine triphosphatase adenosine diphosphate ribosylation factor 6 (ARF6). Binding of WNT5A to the Frizzled 4-LRP6 (low-density lipoprotein receptor-related protein 6) receptor complex activated ARF6, which liberated ß-catenin from N-cadherin, thus increasing the pool of free ß-catenin, enhancing ß-catenin-mediated transcription, and stimulating invasion. In contrast to WNT5A, the guidance cue SLIT2 and its receptor ROBO1 inhibited ARF6 activation and, accordingly, stabilized the interaction of N-cadherin with ß-catenin and reduced transcription and invasion. Thus, ARF6 integrated competing signals in melanoma cells, thereby enabling plasticity in the response to external cues. Moreover, small-molecule inhibition of ARF6 stabilized adherens junctions, blocked ß-catenin signaling and invasiveness of melanoma cells in culture, and reduced spontaneous pulmonary metastasis in mice, suggesting that targeting ARF6 may provide a means of inhibiting WNT/ß-catenin signaling in cancer.
Assuntos
Fatores de Ribosilação do ADP/fisiologia , Melanoma/patologia , Invasividade Neoplásica , Metástase Neoplásica , Proteínas Proto-Oncogênicas/fisiologia , Ativação Transcricional/fisiologia , Proteínas Wnt/fisiologia , beta Catenina/fisiologia , Fator 6 de Ribosilação do ADP , Inativação Gênica , Humanos , Transdução de Sinais , Proteína Wnt-5a , beta Catenina/metabolismoRESUMO
A series of tetrahydropyranyl (THP) derivatives has been developed as potent inhibitors of isoprenylcysteine carboxyl methyltransferase (ICMT) for use as anticancer agents. Structural modification of the submicromolar hit compound 3 led to the potent 3-methoxy substituted analogue 27. Further SAR development around the THP ring resulted in an additional 10-fold increase in potency, exemplified by analogue 75 with an IC(50) of 1.3 nM. Active and potent compounds demonstrated a dose-dependent increase in Ras cytosolic protein. Potent ICMT inhibitors also reduced cell viability in several cancer cell lines with growth inhibition (GI(50)) values ranging from 0.3 to >100 µM. However, none of the cellular effects observed using ICMT inhibitors were as pronounced as those resulting from a farnesyltransferase inhibitor.
Assuntos
Antineoplásicos/síntese química , Proteínas Metiltransferases/antagonistas & inibidores , Piranos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular/efeitos dos fármacos , Citosol/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Técnicas de Inativação de Genes , Humanos , Proteínas Metiltransferases/genética , Piranos/química , Piranos/farmacologia , Proteínas Recombinantes/química , Estereoisomerismo , Relação Estrutura-Atividade , Proteínas ras/biossínteseRESUMO
Mps1 is a dual specificity protein kinase that is essential for the bipolar attachment of chromosomes to the mitotic spindle and for maintaining the spindle assembly checkpoint until all chromosomes are properly attached. Mps1 is expressed at high levels during mitosis and is abundantly expressed in cancer cells. Disruption of Mps1 function induces aneuploidy and cell death. We report the identification of MPI-0479605, a potent and selective ATP competitive inhibitor of Mps1. Cells treated with MPI-0479605 undergo aberrant mitosis, resulting in aneuploidy and formation of micronuclei. In cells with wild-type p53, this promotes the induction of a postmitotic checkpoint characterized by the ATM- and RAD3-related-dependent activation of the p53-p21 pathway. In both wild-type and p53 mutant cells lines, there is a growth arrest and inhibition of DNA synthesis. Subsequently, cells undergo mitotic catastrophe and/or an apoptotic response. In xenograft models, MPI-0479605 inhibits tumor growth, suggesting that drugs targeting Mps1 may have utility as novel cancer therapeutics.
Assuntos
Adenina/análogos & derivados , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Morfolinas/farmacologia , Morfolinas/uso terapêutico , Neoplasias/tratamento farmacológico , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Adenina/isolamento & purificação , Adenina/farmacologia , Adenina/uso terapêutico , Animais , Antineoplásicos/isolamento & purificação , Linhagem Celular Tumoral , Células HCT116 , Humanos , Camundongos , Camundongos Nus , Mitose/efeitos dos fármacos , Mitose/fisiologia , Modelos Biológicos , Peso Molecular , Morfolinas/isolamento & purificação , Neoplasias/patologia , Inibidores de Proteínas Quinases/isolamento & purificação , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Bibliotecas de Moléculas Pequenas/química , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Drug discovery based on cellular phenotypes is impeded by the challenge of identifying the molecular target. To alleviate this problem, we developed a chemical proteomic process to identify cellular proteins that bind to small molecules. CB30865 is a potent (subnanomolar) and selective cytotoxic compound of previously unknown mechanism of action. By combining chemical proteomics with biochemical and cellular pharmacology we have determined that CB30865 cytotoxicity is due to subnanomolar inhibition of nicotinamide phosphoribosyltransferase (Nampt), an enzyme present in the NAD biosynthetic pathway. Cancer cells develop dependence on Nampt due to increased energy requirements and the elevated activity of NAD consuming enzymes such as sirtuins and mono and poly(ADP-ribose) polymerases (PARPs). These findings suggest new chemical starting points for Nampt inhibitors and further implicate this enzyme as a target in cancer.
Assuntos
Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Nicotinamida Fosforribosiltransferase/metabolismo , Produção de Droga sem Interesse Comercial , Proteômica/métodos , Quinazolinas/metabolismo , Quinazolinas/farmacologia , Antineoplásicos/química , Descoberta de Drogas , Células HCT116 , Humanos , Nicotinamida Fosforribosiltransferase/antagonistas & inibidores , Quinazolinas/químicaRESUMO
We have shown previously that the target of the potent cytotoxic agent 4-[(7-bromo-2-methyl-4-oxo-3H-quinazolin-6-yl)methyl-prop-2-ynylamino]-N-(3-pyridylmethyl)benzamide (CB38065, 1) is nicotinamide phosphoribosyltransferase (Nampt). With its cellular target known we sought to optimize the biochemical and cellular Nampt activity of 1 as well as its cytotoxicity. It was found that a 3-pyridylmethylamide substituent in the A region was critical to cellular Nampt activity and cytotoxicity, although other aromatic substitution did yield compounds with submicromolar enzymatic inhibition. Small unsaturated groups worked best in the D-region of the molecule, with 3,3-dimethylallyl providing optimal potency. The E region required a quinazolin-4-one or 1,2,3-benzotriazin-4-one group for activity, and many substituents were tolerated at C² of the quinazolin-4-one. The best compounds showed subnanomolar inhibition of Nampt and low nanomolar cytotoxicity in cellular assays.