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1.
Nat Med ; 25(12): 1938-1947, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31792461

RESUMO

B-cell lymphoma extra large (BCL-XL) is a well-validated cancer target. However, the on-target and dose-limiting thrombocytopenia limits the use of BCL-XL inhibitors, such as ABT263, as safe and effective anticancer agents. To reduce the toxicity of ABT263, we converted it into DT2216, a BCL-XL proteolysis-targeting chimera (PROTAC), that targets BCL-XL to the Von Hippel-Lindau (VHL) E3 ligase for degradation. We found that DT2216 was more potent against various BCL-XL-dependent leukemia and cancer cells but considerably less toxic to platelets than ABT263 in vitro because VHL is poorly expressed in platelets. In vivo, DT2216 effectively inhibits the growth of several xenograft tumors as a single agent or in combination with other chemotherapeutic agents, without causing appreciable thrombocytopenia. These findings demonstrate the potential to use PROTAC technology to reduce on-target drug toxicities and rescue the therapeutic potential of previously undruggable targets. Furthermore, DT2216 may be developed as a safe first-in-class anticancer agent targeting BCL-XL.


Assuntos
Compostos de Anilina/farmacologia , Sulfonamidas/farmacologia , Trombocitopenia/tratamento farmacológico , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Proteína bcl-X/genética , Compostos de Anilina/química , Animais , Antineoplásicos/farmacologia , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Xenoenxertos , Humanos , Camundongos , Proteólise , Sulfonamidas/química , Trombocitopenia/genética , Trombocitopenia/patologia , Proteína bcl-X/antagonistas & inibidores
2.
Chem Commun (Camb) ; 55(98): 14765-14768, 2019 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-31754664

RESUMO

BCL-XL, an anti-apoptotic BCL-2 family protein, plays a key role in cancer cell survival. However, the potential of BCL-XL as an anti-cancer target has been hampered by the on-target platelet toxicity because platelets depend on BCL-XL to maintain their viability. Here we report the development of a PROTAC BCL-XL degrader, XZ424, which has increased selectivity for BCL-XL-dependent MOLT-4 cells over human platelets compared with conventional BCL-XL inhibitors. This proof-of-concept study demonstrates the potential of utilizing a PROTAC approach to achieve tissue selectivity.


Assuntos
Isoquinolinas/química , Talidomida/análogos & derivados , Proteína bcl-X/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Apoptose/efeitos dos fármacos , Plaquetas/citologia , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Linhagem Celular Tumoral , Humanos , Isoquinolinas/metabolismo , Isoquinolinas/farmacologia , Ligação Proteica , Talidomida/química , Talidomida/metabolismo , Talidomida/farmacologia , Proteína bcl-X/antagonistas & inibidores
3.
PLoS One ; 14(5): e0217657, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31150457

RESUMO

Lung cancer is among the common and deadly cancers. Although the treatment options for late-stage cancer patients have continued to increase in numbers, the overall survival rates for these patients have not shown significant improvement. This highlights the need for new targets and drugs to more effectively treat lung cancer patients. In this study, we characterize the MCL-1 inhibitor maritoclax alone or in combination with a BCL-2/xL inhibitor in a panel of lung cancer cell lines. BCL-2 family proteins, phosphorylated proteins, and apoptosis were monitored following the treatments. We found that maritoclax was effective at inhibiting growth in these lung cancer cells. We also establish that cell lines with EGFR mutations were most sensitive to the combined inhibition of MCL-1 and BCL-2/xL. In addition, a high level of phosphorylated AKT (S473) was identified as a marker for sensitivity to the combination treatment. This work has defined EGFR mutations and AKT phosphorylation as markers for sensitivity to combined MCL-1 and BCL-2/xL targeted therapy and establishes a rationale to explore multiple BCL-2 family members in patients who are refractory to EGFR inhibitor treatment. Our data support the design of a clinical trial that aims to employ inhibitors of the BCL-2 family of proteins in lung cancer patients.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Citometria de Fluxo , Humanos , Mutação/genética , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Proteína Oncogênica v-akt/genética , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Pirróis/farmacologia , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/genética
4.
J Recept Signal Transduct Res ; 39(1): 87-97, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31215288

RESUMO

Deregulation of the normal cellular apoptotic function is a fundamental element in the etiology of most cancers and the anti-apoptotic B cell lymphoma 2 (BCL­2) protein family is known to play crucial role in the regulation of this function. Overexpression of this protein family has been implicated in some cancers, such that agents that could inhibit their over-activity are now being explored for anticancer drug development. A number of studies have revealed the anticancer potential of Morinda lucida-derived extracts and compounds. In search of more inhibitors of this anti-apoptotic protein family from plant resources, 47 compounds, identified in Morinda lucida Benth (Rubiaceae) were screened for their inhibitory activities against BCL-XL, BCL-2, and MCL-1 by molecular docking using BINDSURF, while binding interactions of the top compounds were viewed with PyMOL. Druglikeness and Absorption-Distribution-Metabolism-Excretion (ADME) parameters of the top 6 compounds from docking study were evaluated using SuperPred webserver. Results revealed that out of the 47 compounds, 2 triterpenes (ursolic acid and oleanolic acid) and 4 phytosterols (cycloartenol, campesterol, stigmasterol, and ß-sitosterol) have higher binding affinities for the selected BCL-2 proteins, compared to known standard inhibitors; these compounds also fulfill oral drugability of Lipinski rule of five. Therefore, since these Morinda lucida-derived phytosterols and triterpenes show high binding affinity toward the selected anti-apoptotic proteins and exhibited good drugability characteristics, they qualify for further study on drug development against cancers characterized by overexpression of this family of protein.


Assuntos
Morinda/química , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Fitosteróis/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Triterpenos/farmacologia , Proteína bcl-X/antagonistas & inibidores , Simulação por Computador , Humanos , Ligantes , Modelos Moleculares , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Fitosteróis/química , Ligação Proteica , Conformação Proteica , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Triterpenos/química , Proteína bcl-X/metabolismo
5.
Methods Mol Biol ; 1974: 31-40, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31098993

RESUMO

The 2006 discovery of the process of RNA interference opened the door to application of this phenomenon for disease treatments. Short interfering RNA (siRNA) can be used to induce RNA interference, and this approach has generated much interest as a new type of nucleic acid-based drugs in humans and other mammals. However, despite the great potential of siRNA-based drugs in cancer therapy, some drawbacks of siRNAs, such as their instability in vivo and poor cellular uptake, remain unresolved. Here, we review the development of siRNA-based drugs targeting the human Bcl-xL gene, an anti-apoptotic factor overexpressed in many cancers, including prostate cancers. We also introduce a novel application of the biomaterial atelocollagen as a vehicle to functionally deliver tumor-specific siRNA molecules in nude mice. Here, we introduce an orthotopic tumor inoculation model in nude mice: the resulting orthotopic tumors more closely replicate the clinical conditions in humans-including the metastasis mode-than ectopic subcutaneously inoculated tumors.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Neoplasias/genética , RNA Interferente Pequeno/genética , Proteína bcl-X/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Humanos , Camundongos , Neoplasias/terapia , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína bcl-X/antagonistas & inibidores
6.
Cell Oncol (Dordr) ; 42(3): 287-301, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30859392

RESUMO

PURPOSE: Anti-apoptotic and pro-migratory phenotypes are hallmarks of neoplastic diseases, including primary brain malignancies. In this work, we examined whether reprogramming of the apoptotic and migratory machineries through a multi-targeting approach would induce enhanced cell death and enhanced inhibition of the migratory capacity of glioblastoma cells. METHODS: Preclinical testing and molecular analyses of combined inhibition of Bcl-2/Bcl-xL and RAC1 were performed in established, primary cultured and stem-like glioblastoma cell systems. RESULTS: We found that the combined inhibition of Bcl-2/Bcl-xL and RAC1 resulted in synergistic pro-apoptotic and anti-migratory effects in a broad range of different glioblastoma cells. At the molecular level, we found that RAC1 inhibition led to a decreased expression of the deubiquitinase Usp9X, followed by a decreased stability of Mcl-1. We also found that the combined inhibition led to a significantly decreased migratory activity and that tumor formation of glioblastoma cells on chorion allantoic membranes of chicken embryos was markedly impaired following the combined inhibition. CONCLUSIONS: Our data indicate that concomitant inhibition of RAC1 and Bcl-2/Bcl-xL induces pro-apoptotic and anti-migratory glioblastoma phenotypes as well as synergistic anti-neoplastic activities. The clinical efficacy of this inhibitory therapeutic strategy warrants further evaluation.


Assuntos
Compostos de Anilina/farmacologia , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Sulfonamidas/farmacologia , Ubiquitina Tiolesterase/metabolismo , Proteínas rac1 de Ligação ao GTP/antagonistas & inibidores , Antineoplásicos/farmacologia , Apoptose/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Regulação para Baixo/efeitos dos fármacos , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Interferência de RNA , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Ubiquitina Tiolesterase/genética , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/genética , Proteína bcl-X/metabolismo , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo
7.
Methods Mol Biol ; 1877: 351-357, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30536015

RESUMO

The BCL-2 protein family plays central roles in the mitochondrial pathway of cell apoptosis. The BCL-2-Associated X protein (BAX), along with other proapoptotic proteins, induces cell death in response to a variety of stress stimuli. Upon receipt of killing signals, cytosolic BAX is activated and translocates to mitochondria where it causes mitochondrial outer membrane permeabilization (MOMP) and initials a series of cellular events that eventually lead to cell destruction. Despite recent progress toward understanding the structure, function, and activation mechanism of BAX, detailed information about how cytosolic BAX can be inhibited is still limited. Here we describe a method of selecting synthetic antibody fragments (Fabs) against BAX using phage display. Synthetic antibodies discovered from the selection have been used as structural probes to gain novel mechanistic details on BAX inhibition. This synthetic antibody selection method could be potentially applied to other BCL-2 proteins.


Assuntos
Anticorpos/química , Anticorpos/metabolismo , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismo , Apoptose/fisiologia , Citosol/metabolismo , Humanos , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Permeabilidade , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo
8.
ChemMedChem ; 14(1): 100-106, 2019 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-30380184

RESUMO

Progress in drug delivery is hampered by a lack of efficient strategies to target drugs with high specificity and precise spatiotemporal regulation. The remote control of nanoparticles and drugs with light allows regulation of their action site and dosage. Peptide-based drugs are highly specific, non-immunogenic, and can be designed to cross the plasma membrane. In order to combine target specificity and remote control of drug action, here we describe a versatile strategy based on a generalized template to design nanoswitchable peptides that modulate protein-protein interactions upon light activation. This approach is demonstrated to promote photomodulation of two important targets involved in apoptosis (the interactions Bcl-xL-Bak and MDM2-p53), but can be also applied to a large pool of therapeutically relevant protein-protein interactions mediated by α-helical motifs. The template can be adjusted using readily available information about hot spots (residues contributing most to the binding energy) at the protein-protein interface of interest.


Assuntos
Apoptose/efeitos dos fármacos , Nanoestruturas/química , Peptídeos/farmacologia , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Peptídeos/química , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Relação Estrutura-Atividade , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismo , Proteína Killer-Antagonista Homóloga a bcl-2/antagonistas & inibidores , Proteína Killer-Antagonista Homóloga a bcl-2/química , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/química , Proteína bcl-X/metabolismo
9.
Appl Biochem Biotechnol ; 187(3): 1061-1080, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30155742

RESUMO

Selective inhibition is a key focus in the design of chemotherapeutic compounds that can abrogate the oncogenic activities of anti-apoptotic Bcl-2 proteins. Although recent efforts have led to the development of highly selective BH3 mimetics, setbacks such as toxicities have limited their use in cancer therapy. Epigallocatechingallate (EGCG) has been widely reported to selectively inhibit Bcl-2 and Bcl-xL compared to other green tea phenols due to its gallate group. Herein, we investigate the interaction dynamics of EGCG at the hydrophobic grooves of Bcl-2 and Bcl-xL and the consequential effects on their BH4 domains. Arg143 and Asp108 (Bcl-2), and Glu96 and Tyr195 (Bcl-xL) formed high-affinity hydrogen interactions with the gallate group while non-gallate groups of EGCG formed weak interactions. EGCG-bound proteins showed systemic perturbations of BH4 domains coupled with the burial of crucial surface-exposed residues such as Lys17 (Bcl-2) and Asp11 (Bcl-xL); hence, a distortion of non-canonical domain interactions. Interactions of gallate group of EGCG with key hydrophobic groove residues underlie EGCG selectivity while concurrent BH4 domain perturbations potentiate EGCG inhibitory activities. Findings will aid the optimization and design of selective inhibitors that could suppress anti-apoptotic activities of Bcl2-family proteins with minimal toxicities.


Assuntos
Catequina/análogos & derivados , Desenho de Drogas , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sequência de Aminoácidos , Apoptose/efeitos dos fármacos , Catequina/metabolismo , Catequina/farmacologia , Interações Hidrofóbicas e Hidrofílicas , Simulação de Dinâmica Molecular , Domínios Proteicos , Proteínas Proto-Oncogênicas c-bcl-2/química , Especificidade por Substrato , Termodinâmica , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/química , Proteína bcl-X/metabolismo
10.
Cancer Cell ; 34(6): 879-891, 2018 12 10.
Artigo em Inglês | MEDLINE | ID: mdl-30537511

RESUMO

Defects in apoptotic cell death can promote cancer and impair responses of malignant cells to anti-cancer therapy. Pro-survival BCL-2 proteins prevent apoptosis by keeping the cell death effectors, BAX and BAK, in check. The BH3-only proteins initiate apoptosis by neutralizing the pro-survival BCL-2 proteins. Structural analysis and medicinal chemistry led to the development of small-molecule drugs that mimic the function of the BH3-only proteins to kill cancer cells. The BCL-2 inhibitor venetoclax has been approved for treatment of refractory chronic lymphocytic leukemia and this drug and inhibitors of pro-survival MCL-1 and BCL-XL are being tested in diverse malignancies.


Assuntos
Apoptose/efeitos dos fármacos , Materiais Biomiméticos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Sulfonamidas/farmacologia , Antineoplásicos/farmacologia , Humanos , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismo
11.
Eur J Med Chem ; 159: 357-380, 2018 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-30308410

RESUMO

Protein-protein interactions are attractive targets because they control numerous cellular processes. In oncology, apoptosis regulating Bcl-2 family proteins are of particular interest. Apoptotic cell death is controlled via PPIs between the anti-apoptotic proteins hydrophobic groove and the pro-apoptotic proteins BH3 domain. In ovarian carcinoma, it has been previously demonstrated that Bcl-xL and Mcl-1 cooperate to protect tumor cells against apoptosis. Moreover, Mcl-1 is a key regulator of cancer cell survival and is a known resistance factor to Bcl-2/Bcl-xL pharmacological inhibitors making it an attractive therapeutic target. Here, using a structure-guided design from the oligopyridine lead Pyridoclax based on Noxa/Mcl-1 interaction we identified a new derivative, active at lower concentration as compared to Pyridoclax. This new derivative selectively binds to the Mcl-1 hydrophobic groove and releases Bak and Bim from Mcl-1 to induce cell death and sensitize cancer cells to Bcl-2/Bcl-xL targeting strategies.


Assuntos
Desenho de Drogas , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Piridinas/farmacologia , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Interações Hidrofóbicas e Hidrofílicas , Simulação de Dinâmica Molecular , Estrutura Molecular , Proteína de Sequência 1 de Leucemia de Células Mieloides/química , Ligação Proteica , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/química , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade , Células Tumorais Cultivadas , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismo
12.
Sci Rep ; 8(1): 15383, 2018 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-30337641

RESUMO

XPO1 has recently emerged as a viable treatment target for solid malignancies, including glioblastoma (GBM), the most common primary malignant brain tumor in adults. However, given that tumors become commonly resistant to single treatments, the identification of combination therapies is critical. Therefore, we tested the hypothesis that inhibition of anti-apoptotic Bcl-2 family members and XPO1 are synthetically lethal. To this purpose, two clinically validated drug compounds, the BH3-mimetic, ABT263, and the XPO1 inhibitor, Selinexor, were used in preclinical GBM model systems. Our results show that inhibition of XPO1 reduces cellular viability in glioblastoma cell cultures. Moreover, addition of ABT263 significantly enhances the efficacy of XPO1 inhibition on the reduction of cellular viability, which occurs in a synergistic manner. While selinexor inhibits the proliferation of glioblastoma cells, the combination treatment of ABT263 and selinexor results in substantial induction of cell death, which is accompanied by activation of effector- initiator caspases and cleavage of PARP. Mechanistically we find that XPO1 inhibition results in down-regulation of anti-apoptotic Mcl-1 and attenuates ABT263 driven Mcl-1 up-regulation. Consistently, siRNA mediated silencing of Mcl-1 sensitizes for ABT263 mediated cell death and partially for the combination treatment. By using a human patient-derived xenograft model of glioblastoma in mice, we demonstrate that the combination treatment of ABT263 and Selinexor reduces tumor growth significantly more than each compound alone. Collectively, these results suggest that inhibition of XPO1 and Bcl-2/Bcl-xL might be a potential strategy for the treatment of malignant glial tumors.


Assuntos
Compostos de Anilina/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/prevenção & controle , Hidrazinas/farmacologia , Carioferinas/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Sulfonamidas/farmacologia , Triazóis/farmacologia , Proteína bcl-X/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/prevenção & controle , Proliferação de Células/efeitos dos fármacos , Quimioterapia Combinada , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Camundongos , Camundongos Nus , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
13.
PLoS Pathog ; 14(9): e1007299, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30261081

RESUMO

BCL2 family proteins including pro-survival proteins, BH3-only proteins and BAX/BAK proteins control mitochondria-mediated apoptosis to maintain cell homeostasis via the removal of damaged cells and pathogen-infected cells. In this study, we examined the roles of BCL2 proteins in the induction of apoptosis in cells upon infection with flaviviruses, such as Japanese encephalitis virus, Dengue virus and Zika virus. We showed that survival of the infected cells depends on BCLXL, a pro-survival BCL2 protein due to suppression of the expression of another pro-survival protein, MCL1. Treatment with BCLXL inhibitors, as well as deficient BCLXL gene expression, induced BAX/BAK-dependent apoptosis upon infection with flaviviruses. Flavivirus infection attenuates cellular protein synthesis, which confers reduction of short-half-life proteins like MCL1. Inhibition of BCLXL increased phagocytosis of virus-infected cells by macrophages, thereby suppressing viral dissemination and chemokine production. Furthermore, we examined the roles of BCLXL in the death of JEV-infected cells during in vivo infection. Haploinsufficiency of the BCLXL gene, as well as administration of BH3 mimetic compounds, increased survival rate after challenge of JEV infection and suppressed inflammation. These results suggest that BCLXL plays a crucial role in the survival of cells infected with flaviviruses, and that BCLXL may provide a novel antiviral target to suppress propagation of the family of Flaviviridae viruses.


Assuntos
Flavivirus/patogenicidade , Proteína bcl-X/fisiologia , Animais , Apoptose/genética , Apoptose/fisiologia , Linhagem Celular , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Vírus da Dengue/patogenicidade , Vírus da Dengue/fisiologia , Vírus da Encefalite Japonesa (Espécie)/patogenicidade , Vírus da Encefalite Japonesa (Espécie)/fisiologia , Flavivirus/fisiologia , Infecções por Flavivirus/genética , Infecções por Flavivirus/patologia , Infecções por Flavivirus/fisiopatologia , Técnicas de Inativação de Genes , Células HEK293 , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Imunidade Inata , Camundongos , Camundongos Knockout , Modelos Biológicos , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteína de Sequência 1 de Leucemia de Células Mieloides/fisiologia , Células U937 , Células Vero , Replicação Viral/fisiologia , Zika virus/patogenicidade , Zika virus/fisiologia , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/genética
14.
Comput Biol Chem ; 77: 17-27, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30195235

RESUMO

The proteins of Bcl-2 family, which are promising anti-cancer-drug targets, have substantial similarity in primary sequence and share homologous domains as well as similar structural folds. In spite of similarities in sequence and structures, the members of its pro- and anti- apoptotic subgroups form complexes with different type of partners with discriminating binding affinities. Understanding the origin of this discrimination is very important for designing ligands that can either selectively target a protein or could be made broad ranged as necessary. Using principal component analysis (PCA) of the available structures and from the analysis of the evolution of the binding pocket residues, the correlation has been investigated considering two important anti-apoptotic protein Bcl-xl and Mcl-1, which serve as two ideal representatives of this family. The flexibility of the receptor enables them to discriminate between the ligands or the binding partners. It has been observed that although Bcl-xl and Mcl-1 are classified as homologous proteins, through the course of evolution the binding pocket residues are highly conserved for Bcl-xl; whereas they have been substituted frequently in Mcl-1. The investigation has revealed that the Bcl-xl can adjust the backbone conformation of the binding pocket residues to a larger extent to complement with the shape of different binding partners whereas the Mcl-1 shows more variation in the side chain conformation of binding pocket residues for the same purpose.


Assuntos
Proteína de Sequência 1 de Leucemia de Células Mieloides/química , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Proteína bcl-X/química , Proteína bcl-X/metabolismo , Humanos , Ligantes , Modelos Moleculares , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Análise de Componente Principal , Conformação Proteica , Bibliotecas de Moléculas Pequenas/síntese química , Proteína bcl-X/antagonistas & inibidores
15.
Clin Cancer Res ; 24(21): 5458-5470, 2018 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-30021909

RESUMO

Purpose: Clinically available BH3 mimetic drugs targeting BCLXL and/or BCL2 (navitoclax and venetoclax, respectively) are effective in some hematologic malignancies, but have limited efficacy in solid tumors. This study aimed to identify combination therapies that exploit clinical BH3 mimetics for prostate cancer.Experimental Design: Prostate cancer cells or xenografts were treated with BH3 mimetics as single agents or in combination with other agents, and effects on MCL1 and apoptosis were assessed. MCL1 was also targeted directly using RNAi, CRISPR, or an MCL1-specific BH3 mimetic, S63845.Results: We initially found that MCL1 depletion or inhibition markedly sensitized prostate cancer cells to apoptosis mediated by navitoclax, but not venetoclax, in vitro and in vivo, indicating that they are primed to undergo apoptosis and protected by MCL1 and BCLXL. Small-molecule EGFR kinase inhibitors (erlotinib, lapatinib) also dramatically sensitized to navitoclax-mediated apoptosis, and this was associated with markedly increased proteasome-dependent degradation of MCL1. This increased MCL1 degradation appeared to be through a novel mechanism, as it was not dependent upon GSK3ß-mediated phosphorylation and subsequent ubiquitylation by the ubiquitin ligases ßTRCP and FBW7, or through other previously identified MCL1 ubiquitin ligases or deubiquitinases. Inhibitors targeting additional kinases (cabozantinib and sorafenib) similarly caused GSK3ß-independent MCL1 degradation, and in combination with navitoclax drove apoptosis in vitro and in vivo Conclusions: These results show that prostate cancer cells are primed to undergo apoptosis and that cotargeting BCLXL and MCL1, directly or indirectly through agents that increase MCL1 degradation, can induce dramatic apoptotic responses. Clin Cancer Res; 24(21); 5458-70. ©2018 AACR.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Neoplasias da Próstata/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteína bcl-X/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Técnicas de Silenciamento de Genes , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Masculino , Camundongos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Proteólise , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Chemistry ; 24(52): 13762-13766, 2018 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-29917279

RESUMO

Strain-promoted azide-alkyne cycloadditions (SPAAC) have proven extremely useful for labeling of biomolecules, but typically produce isomeric mixtures. This is not appropriate for the formation of bioactive molecules in living cells. Here, the first use of SPAAC for the isomer-free synthesis of a bioactive molecule is reported both in vitro and inside cultured cells. We developed the symmetrical cyclooctyne SYPCO and used it for the generation of a chemically uniform triazole inhibitor of protein-protein interactions mediated by Bcl-xL via isomer-free SPAAC (iSPAAC). Tumor cells treated with the reactants of the iSPAAC reaction contained higher concentrations of triazole, and displayed higher apoptosis levels, than cells treated with pre-synthesized triazole. We envision iSPAAC as a broadly applicable method for modulating intracellular targets with organic molecules with molecular weights prohibitively large for cellular uptake, via smaller and thus more cell-permeable components.


Assuntos
Antineoplásicos/síntese química , Triazóis/síntese química , Proteína bcl-X/antagonistas & inibidores , Alquinos/química , Alquinos/farmacologia , Antineoplásicos/farmacologia , Apoptose , Azidas/química , Azidas/farmacologia , Reação de Cicloadição , Humanos , Isomerismo , Células K562 , Cinética , Simulação de Acoplamento Molecular , Peso Molecular , Ligação Proteica , Triazóis/farmacologia
17.
Life Sci ; 205: 9-17, 2018 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-29738779

RESUMO

AIMS: As a fifth most common cancer type, Hepatocellularcarcinoma (HCC) ranked third leading cause of cancer deaths worldwide. Arsenic trioxide (As2O3) is known as chemotherapeutic agent against few cancer including Acute promyelocyticleukemia and solid tumors. But its effect and possible associated mechanism in HCC is meager. Present study aimed to assess As2O3 modulatory effect on liver cancer by assessing cell growth and viability. METHODS: Liver normal (Chang liver) and cancerous cells (Hep3B) were exposed to different concentration's (0, 1, 5, 10 & 15 µM) of As2O3 at different intervals (24, 48 & 72 h). Cell growth was assessed microscopically, and Cytotoxicity assays were done through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Water-soluble tetrazolium salt (WST) growth inhibition assays. Cell viability was studied by trypan blue staining. Apoptosis was analyzed by Annexin V/PI assay, and expression of genes (Notch and anti-apoptotic) were determined through western blotting and Q-PCR method. KEY FINDINGS: A significant reduction in cell growth and viability was reported in liver cancerous cells as compare to normal cells at 5 µM As2O3. Consistently, As2O3 induced apoptosis along with down-regulation of anti-apoptotic protein Bcl-xL, and up regulates expression of Notch that leads towards apoptosis. SIGNIFICANCE: Results clearly suggest that As2O3 restricted growth and induces apoptosis more in liver cancer cells as compared to normal cells. This finding suggests that it could be a promising potential therapeutic agent against liver cancer which need further testing by in-vivo investigations.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Arsenicais/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Óxidos/farmacologia , Trióxido de Arsênio , Carcinoma Hepatocelular/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Humanos , Neoplasias Hepáticas/patologia , Receptores Notch/biossíntese , Receptores Notch/efeitos dos fármacos , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/biossíntese
18.
Sci Rep ; 8(1): 7373, 2018 05 09.
Artigo em Inglês | MEDLINE | ID: mdl-29743557

RESUMO

Recent data suggest that glioblastomas (GBM) activate the c-MET signaling pathway and display increased levels in anti-apoptotic Bcl-2 family members. Therefore, targeting these two deregulated pathways for therapy might yield synergistic treatment responses. We applied extracellular flux analysis to assess tumor metabolism. We found that combined treatment with ABT263 and Crizotinib synergistically reduces the proliferation of glioblastoma cells, which was dependent on dual inhibition of Bcl-2 and Bcl-xL. The combination treatment led to enhanced apoptosis with loss of mitochondrial membrane potential and activation of caspases. On the molecular level, c-MET-inhibition results in significant energy deprivation with a reduction in oxidative phosphorylation, respiratory capacity and a suppression of intracellular energy production (ATP). In turn, loss of energy levels suppresses protein synthesis, causing a decline in anti-apoptotic Mcl-1 levels. Silencing of Mcl-1 enhanced ABT263 and MET-inhibitor mediated apoptosis, but marginally the combination treatment, indicating that Mcl-1 is the central factor for the induction of cell death induced by the combination treatment. Finally, combined treatment with BH3-mimetics and c-MET inhibitors results in significantly smaller tumors than each treatment alone in a PDX model system of glioblastoma. These results suggest that c-MET inhibition causes a selective vulnerability of GBM cells to Bcl-2/Bcl-xL inhibition.


Assuntos
Compostos de Anilina/farmacologia , Glioblastoma/patologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Sulfonamidas/farmacologia , Proteína bcl-X/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Crizotinibe/farmacologia , Sinergismo Farmacológico , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Humanos , Camundongos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Oncogene ; 37(32): 4475-4488, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29743589

RESUMO

Genetic alterations in the fibroblast growth factor receptors (FGFRs) have been described in multiple solid tumours including bladder cancer, head and neck and lung squamous cell carcinoma (SqCC). However, recent clinical trials showed limited efficacy of FGFR-targeted therapy in lung SqCC, suggesting combination therapy may be necessary to improve patient outcomes. Here we demonstrate that FGFR therapy primes SqCC for cell death by increasing the expression of the pro-apoptotic protein BIM. We therefore hypothesised that combining BH3-mimetics, potent inhibitors of pro-survival proteins, with FGFR-targeted therapy may enhance the killing of SqCC cells. Using patient-derived xenografts and specific inhibitors of BCL-2, BCL-XL, and MCL-1, we identified a greater reliance of lung SqCC cells on BCL-XL and MCL-1 compared to BCL-2 for survival. However, neither BCL-XL nor MCL-1 inhibitors alone provided a survival benefit in combination FGFR therapy in vivo. Only triple BCL-XL, MCL-1, and FGFR inhibition resulted in tumour volume regression and prolonged survival in vivo, demonstrating the ability of BCL-XL and MCL-1 proteins to compensate for each other in lung SqCC. Our work therefore provides a rationale for the inhibition of MCL-1, BCL-XL, and FGFR1 to maximize therapeutic response in FGFR1-expressing lung SqCC.


Assuntos
Antineoplásicos/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Receptores de Fatores de Crescimento de Fibroblastos/antagonistas & inibidores , Proteína bcl-X/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/metabolismo , Proteína 11 Semelhante a Bcl-2/metabolismo , Carcinoma de Células Escamosas/metabolismo , Linhagem Celular Tumoral , Quimioterapia Combinada/métodos , Humanos , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
20.
Bioorg Med Chem Lett ; 28(7): 1171-1177, 2018 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-29534928

RESUMO

For many years, studies focused on developing new natural or synthetic compounds with antineoplastic activity have attracted the attention of researchers. An interesting group of such compounds seem to be those with both lactone moiety and an aromatic ring which, in addition to antimicrobial or antiviral activity, also exhibit antitumor properties. The study shows antitumor activity of two enantiomeric trans isomers of 5-(1-iodoethyl)-4-(2',5'-dimethylphenyl)dihydrofuran-2-one. Our aim was to determine their antitumor activity manifested as an ability to induce apoptosis in selected canine cancer cell lines as well as to evaluate differences in their strength depending on the configuration of their stereogenic centers. The enantiomers (+)-(4R,5S,6R)-1 and (-)-(4S,5R,6S)-2 were found to induce classical caspase-dependent apoptosis through downregulation of the expression of anti-apoptotic proteins Bcl-xL and Bcl-2. Although the mechanism of apoptosis induction was the same for both enantiomers, they differed in their strength, as stronger antineoplastic activity in vitro was exhibited by isomer (+)-(4R,5S,6R)-1.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzaldeídos/farmacologia , Regulação para Baixo/efeitos dos fármacos , Lactonas/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteína bcl-X/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Cães , Relação Dose-Resposta a Droga , Lactonas/síntese química , Lactonas/química , Linfoma/tratamento farmacológico , Linfoma/patologia , Estrutura Molecular , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Estereoisomerismo , Relação Estrutura-Atividade , Proteína bcl-X/metabolismo
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