RESUMO
Upregulation of antiapoptotic Bcl-2 proteins in certain tumors confers cancer cell resistance to chemotherapy or radiations. Members of the antiapoptotic Bcl-2 proteins, including Bcl-2, Mcl-1, Bcl-xL, Bcl-w, and Bfl-1, inhibit apoptosis by selectively binding to conserved α-helical regions, named BH3 domains, of pro-apoptotic proteins such as Bim, tBid, Bad, or NOXA. Five antiapoptotic proteins have been identified that interact with various selectivity with BH3 containing pro-apoptotic counterparts. Cancer cells present various and variable levels of these proteins, making the design of effective apoptosis based therapeutics challenging. Recently, BH3 profiling was introduced as a method to classify cancer cells based on their ability to resist apoptosis following exposure to selected BH3 peptides. However, these studies were based on binding affinities measured with model BH3 peptides and Bcl-2-proteins taken from mouse sequences. While the majority of these interactions are conserved between mice and humans, we found surprisingly that human NOXA binds to human Bfl-1 potently and covalently via conserved Cys residues, with over 2 orders of magnitude increased affinity over hMcl-1. Our data suggest that some assumptions of the original BH3 profiling need to be revisited and that perhaps further targeting efforts should be redirected toward Bfl-1, for which no suitable specific inhibitors or pharmacological tools have been reported. In this regard, we also describe the initial design and characterizations of novel covalent BH3-based agents that potently target Bfl-1. These molecules could provide a novel platform on which to design effective Bfl-1 targeting therapeutics.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos , Antígenos de Histocompatibilidade Menor/metabolismo , Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sequência de Aminoácidos , Apoptose , Linhagem Celular Tumoral , HumanosRESUMO
The cell surface receptor CD44 is a glycoprotein belonging to the hyaluronan-binding proteins, termed hyaladherins. CD44 is expressed in a wide variety of isoforms in many cells and, in particular, is present on the surface of malignant cells where it is involved in the onset and progression of cancer. In a first attempt to identify novel CD44-binding agents, we first characterized, with NMR spectroscopic techniques, several agents that were reported to bind to human CD44 (hCD44). To our surprise, however, none of these putative CD44-binding agents, including a peptide that is in phaseâ 2 clinical trials (A6 peptide) and a recently reported fragment hit, were found to interact significantly with recombinant hCD44(21-178). Nonetheless, we further report that a fragment-screening campaign, with solution NMR spectroscopy as the detection method, identified a viable fragment hit that bound in a potentially functional pocket on the surface of CD44, opposite to the hyaluronic acid binding site. We hypothesize that this pocket could be indirectly associated with the cellular and inâ vivo activity of the A6 peptide, which would provide a novel framework for the possible development of therapeutically viable CD44 antagonists.
Assuntos
Receptores de Hialuronatos/metabolismo , Ligantes , Peptídeos/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Calorimetria , Humanos , Receptores de Hialuronatos/química , Receptores de Hialuronatos/genética , Ressonância Magnética Nuclear Biomolecular , Peptídeos/química , Ligação Proteica , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificaçãoRESUMO
First line treatment for pancreatic cancer consists of surgical resection, if possible, and a subsequent course of chemotherapy using the nucleoside analogue gemcitabine. In some patients, an active transport mechanism allows gemcitabine to enter efficiently into the tumor cells, resulting in a significant clinical benefit. However, in most patients, low expression of gemcitabine transporters limits the efficacy of the drug to marginal levels, and patients need frequent administration of the drug at high doses, significantly increasing systemic drug toxicity. In this article we focus on a novel targeted delivery approach for gemcitabine consisting of conjugating the drug with an EphA2 targeting agent. We show that the EphA2 receptor is highly expressed in pancreatic cancers, and accordingly, the drug-conjugate is more effective than gemcitabine alone in targeting pancreatic tumors. Our preliminary observations suggest that this approach may provide a general benefit to pancreatic cancer patients and offers a comprehensive strategy for enhancing delivery of diverse therapeutic agents to a wide range of cancers overexpressing EphA2, thereby potentially reducing toxicity while enhancing therapeutic efficacy.
Assuntos
Antimetabólitos Antineoplásicos/administração & dosagem , Desoxicitidina/análogos & derivados , Terapia de Alvo Molecular/métodos , Neoplasias Pancreáticas/tratamento farmacológico , Receptor EphA2/metabolismo , Animais , Linhagem Celular Tumoral , Desoxicitidina/administração & dosagem , Humanos , Camundongos , Camundongos Nus , Ensaios Antitumorais Modelo de Xenoenxerto , GencitabinaRESUMO
Disrupting the eukaryotic translation initiation factor 4F (eIF4F) complex offers an appealing strategy to potentiate the effectiveness of existing cancer therapies and to overcome resistance to drugs such as BRAF inhibitors (BRAFi). Here, we identified and characterized the small molecule SBI-0640756 (SBI-756), a first-in-class inhibitor that targets eIF4G1 and disrupts the eIF4F complex. SBI-756 impaired the eIF4F complex assembly independently of mTOR and attenuated growth of BRAF-resistant and BRAF-independent melanomas. SBI-756 also suppressed AKT and NF-κB signaling, but small-molecule derivatives were identified that only marginally affected these pathways while still inhibiting eIF4F complex formation and melanoma growth, illustrating the potential for further structural and functional manipulation of SBI-756 as a drug lead. In the gene expression signature patterns elicited by SBI-756, DNA damage, and cell-cycle regulatory factors were prominent, with mutations in melanoma cells affecting these pathways conferring drug resistance. SBI-756 inhibited the growth of NRAS, BRAF, and NF1-mutant melanomas in vitro and delayed the onset and reduced the incidence of Nras/Ink4a melanomas in vivo. Furthermore, combining SBI-756 and a BRAFi attenuated the formation of BRAFi-resistant human tumors. Taken together, our findings show how SBI-756 abrogates the growth of BRAF-independent and BRAFi-resistant melanomas, offering a preclinical rationale to evaluate its antitumor effects in other cancers.
Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Fator de Iniciação 4F em Eucariotos/metabolismo , Lactamas/farmacologia , Melanoma/patologia , Quinolonas/farmacologia , Animais , Western Blotting , Linhagem Celular Tumoral , Modelos Animais de Doenças , Técnicas de Inativação de Genes , Humanos , Melanoma/metabolismo , CamundongosRESUMO
The development of novel, targeted delivery agents for anti-cancer therapies requires the design and optimization of potent and selective tumor-targeting agents that are stable and amenable to conjugation with chemotherapeutic drugs. While short peptides represent potentially an excellent platform for these purposes, they often get degraded and are eliminated too rapidly in vivo. In this study, we used a combination of nuclear magnetic resonance-guided structure-activity relationships along with biochemical and cellular studies to derive a novel tumor-homing agent, named 123B9, targeting the EphA2 tyrosine kinase receptor ligand-binding domain. Conjugating 123B9 to the chemotherapeutic drug paclitaxel (PTX) via a stable linker results in an agent that is significantly more effective than the unconjugated drug in both a pancreatic cancer xenograft model and a melanoma lung colonization and metastases model. Hence, 123B9 could represent a promising strategy for the development of novel targeted therapies for cancer.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Paclitaxel/análogos & derivados , Receptor EphA2/agonistas , Sequência de Aminoácidos , Animais , Antineoplásicos/síntese química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sistemas de Liberação de Medicamentos/métodos , Feminino , Humanos , Masculino , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/metabolismo , Camundongos , Camundongos Nus , Modelos Animais , Terapia de Alvo Molecular , Paclitaxel/química , Paclitaxel/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/metabolismo , Ratos , Receptor EphA2/química , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
In recent years the ever so complex field of drug discovery has embraced novel design strategies based on biophysical fragment screening (fragment-based drug design; FBDD) using nuclear magnetic resonance spectroscopy (NMR) and/or structure-guided approaches, most often using X-ray crystallography and computer modeling. Experience from recent years unveiled that these methods are more effective and less prone to artifacts compared to biochemical high-throughput screening (HTS) of large collection of compounds in designing protein inhibitors. Hence these strategies are increasingly becoming the most utilized in the modern pharmaceutical industry. Nonetheless, there is still an impending need to develop innovative and effective strategies to tackle other more challenging targets such as those involving protein-protein interactions (PPIs). While HTS strategies notoriously fail to identify viable hits against such targets, few successful examples of PPIs antagonists derived by FBDD strategies exist. Recently, we reported on a new strategy that combines some of the basic principles of fragment-based screening with combinatorial chemistry and NMR-based screening. The approach, termed HTS by NMR, combines the advantages of combinatorial chemistry and NMR-based screening to rapidly and unambiguously identify bona fide inhibitors of PPIs. This review will reiterate the critical aspects of the approach with examples of possible applications.
Assuntos
Descoberta de Drogas , Proteína de Sequência 1 de Leucemia de Células Mieloides/química , Receptor EphA4/química , Bibliotecas de Moléculas Pequenas/química , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/química , Técnicas de Química Combinatória , Cristalografia por Raios X , Efrina-A5/química , Ensaios de Triagem em Larga Escala , Humanos , Ligantes , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Peptídeos/química , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas , Receptor EphA4/antagonistas & inibidores , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/antagonistas & inibidoresRESUMO
From the whole plant of Euphorbia cyparissias, two new diterpenes based on jatrophane skeleton, named cyparissins A and B (1 and 2) were isolated. Their chemical structures were established through a combination of nuclear magnetic resonance spectroscopy and mass spectrometric methods. The new cyparissins A and B were tested to evaluate their ability to inhibit P-glycoprotein-mediated multidrug resistance and their cytotoxic activity against A2780 human ovarian cancer cells, both WT and ADR. Compounds 1 and 2 showed moderate inhibitory effects on P-glycoprotein while showing a significant concentration-depending cytotoxic activity for both cancer cell lines. These isolated compounds are based on a new chemical structure that expands the knowledge base for this class of bioactive metabolites.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Diterpenos/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Euphorbia/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/antagonistas & inibidores , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Linhagem Celular Tumoral/efeitos dos fármacos , Feminino , Humanos , Estrutura Molecular , Neoplasias Ovarianas/patologiaRESUMO
Because of its overexpression in a range of solid tumors, the EphA2 receptor is a validated target for cancer therapeutics. We recently described a new targeted delivery system based on specific EphA2-targeting peptides conjugated with the chemotherapeutic agent paclitaxel. Here, we investigate the chemical determinants responsible for the stability and degradation of these agents in plasma. Introducing modifications in both the peptide and the linker between the peptide and paclitaxel resulted in drug conjugates that are both long-lived in rat plasma and that markedly decrease tumor size in a prostate cancer xenograft model compared with paclitaxel alone treatment. These studies identify critical rate-limiting degradation sites on the peptide-drug conjugates, enabling the design of agents with increased stability and efficacy. These results provide support for our central hypothesis that peptide-drug conjugates targeting EphA2 represent an innovative and potentially effective strategy to selectively deliver cytotoxic drugs to cancer cells.
Assuntos
Portadores de Fármacos/síntese química , Desenho de Fármacos , Receptor EphA2/metabolismo , Sequência de Aminoácidos , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Portadores de Fármacos/química , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Paclitaxel/química , Paclitaxel/farmacologia , Paclitaxel/uso terapêutico , Peptídeos/síntese química , Peptídeos/química , Neoplasias da Próstata/tratamento farmacológico , Receptor EphA2/química , Transplante HeterólogoRESUMO
The protein Kinase B alpha (AKT) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways are central regulators of cellular signaling events at the basis of tumor development and progression. Both pathways are often up-regulated in different tumor types including melanoma. We recently reported the identification of compound 1 (BI-69A11) as inhibitor of the AKT and the NF-κB pathways. Here, we describe SAR studies that led to novel fluorinated derivatives with increased cellular potency, reflected in efficient inhibition of AKT and IKKs. Selected compounds demonstrated effective toxicity on melanoma, breast, and prostate cell lines. Finally, a representative derivative showed promising efficacy in an in vivo melanoma xenograft model.
Assuntos
NF-kappa B/antagonistas & inibidores , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Animais , Benzimidazóis/química , Benzimidazóis/farmacologia , Benzimidazóis/toxicidade , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Feminino , Humanos , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Camundongos Nus , NF-kappa B/metabolismo , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/toxicidade , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinolonas/química , Quinolonas/farmacologia , Quinolonas/toxicidade , Relação Estrutura-Atividade , Transplante HeterólogoRESUMO
To date, there are no effective therapies for tumors bearing NRAS mutations, which are present in 15-20% of human melanomas. Here we extend our earlier studies where we demonstrated that the small molecule BI-69A11 inhibits the growth of melanoma cell lines. Gene expression analysis revealed the induction of interferon- and cell death-related genes that were associated with responsiveness of melanoma cell lines to BI-69A11. Strikingly, the administration of BI-69A11 inhibited melanoma development in genetically modified mice bearing an inducible form of activated Nras and a deletion of the Ink4a gene (Nras((Q61K)) ::Ink4a(-/-) ). Biweekly administration of BI-69A11 starting at 10 weeks or as late as 24 weeks after the induction of mutant Nras expression inhibited melanoma development (100 and 36%, respectively). BI-69A11 treatment did not inhibit the development of histiocytic sarcomas, which constitute about 50% of the tumors in this model. BI-69A11-resistant Nras((Q61K)) ::Ink4a(-/-) tumors exhibited increased CD45 expression, reflective of immune cell infiltration and upregulation of gene networks associated with the cytoskeleton, DNA damage response, and small molecule transport. The ability to attenuate the development of NRAS mutant melanomas supports further development of BI-69A11 for clinical assessment.
Assuntos
Substituição de Aminoácidos/genética , Benzimidazóis/uso terapêutico , Inibidor p16 de Quinase Dependente de Ciclina/deficiência , Melanoma/tratamento farmacológico , Lesões Pré-Cancerosas/tratamento farmacológico , Quinolonas/uso terapêutico , Neoplasias Cutâneas/tratamento farmacológico , Proteínas ras/genética , Animais , Benzimidazóis/administração & dosagem , Benzimidazóis/farmacologia , Proliferação de Células/efeitos dos fármacos , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Injeções Intraperitoneais , Antígeno Ki-67/metabolismo , Antígenos Comuns de Leucócito/metabolismo , Melanoma/genética , Melanoma/patologia , Camundongos , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/patologia , Proteínas Proto-Oncogênicas B-raf/genética , Quinolonas/administração & dosagem , Quinolonas/farmacologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/uso terapêutico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PDK1 is a key member of the AGC protein kinase family. It plays an important role in a variety of cellular functions, leading to the activation of the PI3K signaling pathway, an event often associated with the onset and progression of several human cancers. Numerous recent observations suggest that PDK1 inhibitors may provide novel opportunities for the development of effective classes of therapeutics. On these premises, recent years have witnessed an increased effort by medicinal chemists to develop novel scaffolds to derive potent and selective PDK1 inhibitors. The intent of this review is to update the reader on the recent patent literature, covering applications published between June 2008 and September 2011 that report on PDK1 inhibitors.
Assuntos
Proteínas Quinases Dependentes de 3-Fosfoinositídeo/antagonistas & inibidores , Desenho de Fármacos , Inibidores de Proteínas Quinases/farmacologia , Animais , Antineoplásicos/farmacologia , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Patentes como Assunto , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/fisiologiaRESUMO
In melanoma, the activation of pro-survival signaling pathways, such as the AKT and NF-κB pathways, is critical for tumor growth. We have recently reported that the AKT inhibitor BI-69A11 causes efficient inhibition of melanoma growth. Here, we show that in addition to its AKT inhibitory activity, BI-69A11 also targets the NF-κB pathway. In melanoma cell lines, BI-69A11 inhibited TNF-α-stimulated IKKα/ß and IκB phosphorylation as well as NF-κB reporter gene expression. Furthermore, the effective inhibition of melanoma growth by BI-69A11 was attenuated upon NF-κB activation. Mechanistically, reduced NF-κB signaling by BI-69-A11 is mediated by the inhibition of sphingosine kinase 1, identified in a screen of 315 kinases. Significantly, we demonstrate that BI-69A11 is well tolerated and orally active against UACC 903 and SW1 melanoma xenografts. Our results demonstrate that BI-69A11 inhibits both the AKT and the NF-κB pathways and that the dual targeting of these pathways may be efficacious as a therapeutic strategy in melanoma.
Assuntos
Benzimidazóis/uso terapêutico , Melanoma/tratamento farmacológico , Terapia de Alvo Molecular , NF-kappa B/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Quinolonas/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Neoplasias Cutâneas/tratamento farmacológico , Administração Oral , Animais , Benzimidazóis/administração & dosagem , Benzimidazóis/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Melanoma/enzimologia , Camundongos , Camundongos Nus , NF-kappa B/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinolonas/administração & dosagem , Quinolonas/farmacologia , Neoplasias Cutâneas/enzimologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
We report comprehensive structure-activity relationship studies on a novel series of c-Jun N-terminal kinase (JNK) inhibitors. Intriguingly, the compounds have a dual inhibitory activity by functioning as both ATP and JIP mimetics, possibly by binding to both the ATP binding site and to the docking site of the kinase. Several of such novel compounds display potent JNK inhibitory profiles both in vitro and in cell.
Assuntos
Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/síntese química , Tiofenos/farmacologia , Trifosfato de Adenosina , Sítios de Ligação , Linhagem Celular , Desenho de Fármacos , Humanos , Mimetismo Molecular , Ligação Proteica , Inibidores de Proteínas Quinases/farmacologia , Relação Estrutura-Atividade , Tiofenos/químicaRESUMO
Our focus in the past several years has been on the identification of novel and effective pan-Bcl-2 antagonists. We have recently reported a series of Apogossypolone (ApoG2) derivatives, resulting in the chiral compound (±) BI97D6. We report here the synthesis and evaluation on its optically pure (-) and (+) atropisomers. Compound (-) BI97D6 potently inhibits the binding of BH3 peptides to Bcl-X(L), Bcl-2, Mcl-1, and Bfl-1 with IC(50) values of 76 ± 5, 31 ± 2, 25 ± 8, and 122 ± 28 nM, respectively. In a cellular assay, compound (-) BI97D6 effectively inhibits cell growth in the PC-3 human prostate cancer and H23 human lung cancer cell lines with EC(50) values of 0.22 ± 0.08 and 0.14 ± 0.02 µM, respectively. Similarly, compound (-) BI97D6 effectively induces apoptosis in the BP3 human lymphoma cell line in a dose-dependent manner. The compound also shows little cytotoxicity against bax(-/-)/bak(-/-) cells, suggesting that it kills cancers cells predominantly via a Bcl-2 pathway. Moreover, compound (-) BI97D6 displays in vivo efficacy in both a Bcl-2-transgenic mouse model and in a prostate cancer xenograft model in mice. Therefore, compound (-) BI97D6 represents a promising drug lead for the development of novel apoptosis-based therapies for cancer.
RESUMO
A new series of 3-ethynyl-1H-indazoles has been synthesized and evaluated in both biochemical and cell-based assays as potential kinase inhibitors. Interestingly, a selected group of compounds identified from this series exhibited low micromolar inhibition against critical components of the PI3K pathway, targeting PI3K, PDK1, and mTOR kinases. A combination of computational modeling and structure-activity relationship studies reveals a possible novel mode for PI3K inhibition, resulting in a PI3Kα isoform-specific compound. Hence, by targeting the most oncogenic mutant isoform of PI3K, the compound displays antiproliferative activity both in monolayer human cancer cell cultures and in three-dimensional tumor models. Because of its favorable physicochemical, in vitro ADME and drug-like properties, we propose that this novel ATP mimetic scaffold could prove useful in deriving novel selecting and multikinase inhibitors for clinical use.