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1.
J Immunol ; 208(5): 1115-1127, 2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-35165166

RESUMO

Purinergic signaling plays a major role in T cell activation leading to IL-2 production and proliferation. However, it is unclear whether purinergic signaling contributes to the differentiation and activation of effector T cells. In this study, we found that the purinergic receptor P2X4 was associated with human Th17 cells but not with Th1 cells. Inhibition of P2X4 receptor with the specific antagonist 5-BDBD and small interfering RNA inhibited the development of Th17 cells and the production of IL-17 by effector Th17 cells stimulated via the CD3/CD28 pathway. Our results showed that P2X4 was required for the expression of retinoic acid-related orphan receptor C, which is the master regulator of Th17 cells. In contrast, inhibition of P2X4 receptor had no effect on Th1 cells and on the production of IFN-γ and it did not affect the expression of the transcription factor T-bet (T-box transcription factor). Furthermore, inhibition of P2X4 receptor reduced the production of IL-17 but not of IFN-γ by effector/memory CD4+ T cells isolated from patients with rheumatoid arthritis. In contrast to P2X4, inhibition of P2X7 and P2Y11 receptors had no effects on Th17 and Th1 cell activation. Finally, treatment with the P2X4 receptor antagonist 5-BDBD reduced the severity of collagen-induced arthritis in mice by inhibiting Th17 cell expansion and activation. Our findings provide novel insights into the role of purinergic signaling in T cell activation and identify a critical role for the purinergic receptor P2X4 in Th17 activation and in autoimmune arthritis.


Assuntos
Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/imunologia , Antagonistas do Receptor Purinérgico P2X/farmacologia , Receptores Purinérgicos P2X4/metabolismo , Células Th17/imunologia , Animais , Artrite Reumatoide/patologia , Benzodiazepinonas/farmacologia , Diferenciação Celular/imunologia , Células Cultivadas , Humanos , Memória Imunológica/imunologia , Interferon gama/biossíntese , Interleucina-17/biossíntese , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Endogâmicos DBA , Receptores Nucleares Órfãos , Interferência de RNA , RNA Interferente Pequeno/genética , Receptores Purinérgicos P2X4/genética , Proteínas com Domínio T/biossíntese , Células Th1/citologia , Células Th1/imunologia , Células Th17/citologia
2.
Mol Pharmacol ; 101(1): 33-44, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34718224

RESUMO

The P2X4 receptor is a ligand-gated ion channel activated by extracellular ATP. P2X4 activity is associated with neuropathic pain, vasodilation, and pulmonary secretion and is therefore of therapeutic interest. The structure-activity relationship of P2X4 antagonists is poorly understood. Here we elucidate the structure-activity of 5-(3-bromophenyl)-1,3-dihydro-2H-benzofuro[3,2-e]-1,4-diazepin-2-one (5-BDBD) at human P2X4 by combining pharmacology, electrophysiology, molecular modeling, and medicinal chemistry. 5-BDBD antagonized P2X4 in a noncompetitive manner but lacked effect at human P2X2. Molecular modeling and site-directed mutagenesis suggested an allosteric binding site for 5-BDBD located between two subunits in the body region of P2X4, with M109, F178, Y300, and I312 on one subunit and R301 on the neighboring subunit as key residues involved in antagonist binding. The bromine group of 5-BDBD was redundant for the antagonist activity of 5-BDBD, although an interaction between the carbonyl group of 5-BDBD and R301 in P2X4 was associated with 5-BDBD activity. 5-BDBD could inhibit the closed channel but poorly inhibited the channel in the open/desensitizing state. We hypothesize that this is due to constriction of the allosteric site after transition from closed to open channel state. We propose that M109, F178, Y300, R301, and I312 are key residues for 5-BDBD binding; provide a structural explanation of how they contribute to 5-BDBD antagonism; and highlight that the limited action of 5-BDBD on open versus closed channels is due to a conformational change in the allosteric site. SIGNIFICANCE STATEMENT: Activity of P2X4 receptor is associated with neuropathic pain, inflammation, and vasodilatation. Molecular information regarding small-molecule interaction with P2X4 is very limited. Here, this study provides a structural explanation for the action of the small-molecule antagonist 5-BDBD at the human P2X4 receptor.


Assuntos
Benzodiazepinonas/química , Benzodiazepinonas/metabolismo , Antagonistas do Receptor Purinérgico P2X/química , Antagonistas do Receptor Purinérgico P2X/metabolismo , Receptores Purinérgicos P2X4/química , Receptores Purinérgicos P2X4/metabolismo , Regulação Alostérica/efeitos dos fármacos , Regulação Alostérica/fisiologia , Benzodiazepinonas/farmacologia , Células HEK293 , Humanos , Simulação de Dinâmica Molecular , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Antagonistas do Receptor Purinérgico P2X/farmacologia
3.
J Pathol ; 256(2): 149-163, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34652816

RESUMO

Prostate cancer (PCa) remains a leading cause of cancer-related deaths in American men and treatment options for metastatic PCa are limited. There is a critical need to identify new mechanisms that contribute to PCa progression, that distinguish benign from lethal disease, and that have potential for therapeutic targeting. P2X4 belongs to the P2 purinergic receptor family that is commonly upregulated in cancer and is associated with poorer outcomes. We observed P2X4 protein expression primarily in epithelial cells of the prostate, a subset of CD66+ neutrophils, and most CD68+ macrophages. Our analysis of tissue microarrays representing 491 PCa cases demonstrated significantly elevated P2X4 expression in cancer- compared with benign-tissue spots, in prostatic intraepithelial neoplasia, and in PCa with ERG positivity or with PTEN loss. High-level P2X4 expression in benign tissues was likewise associated with the development of metastasis after radical prostatectomy. Treatment with the P2X4-specific agonist cytidine 5'-triphosphate (CTP) increased Transwell migration and invasion of PC3, DU145, and CWR22Rv1 PCa cells. The P2X4 antagonist 5-(3-bromophenyl)-1,3-dihydro-2H-benzofuro[3,2-e]-1,4-diazepin-2-one (5-BDBD) resulted in a dose-dependent decrease in viability of PC3, DU145, LNCaP, CWR22Rv1, TRAMP-C2, Myc-CaP, BMPC1, and BMPC2 cells and decreased DU145 cell migration and invasion. Knockdown of P2X4 attenuated growth, migration, and invasion of PCa cells. Finally, knockdown of P2X4 in Myc-CaP cells resulted in significantly attenuated subcutaneous allograft growth in FVB/NJ mice. Collectively, these data strongly support a role for the P2X4 purinergic receptor in PCa aggressiveness and identify P2X4 as a candidate for therapeutic targeting. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Assuntos
Antineoplásicos/farmacologia , Benzodiazepinonas/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias da Próstata/tratamento farmacológico , Antagonistas do Receptor Purinérgico P2X/farmacologia , Receptores Purinérgicos P2X4/efeitos dos fármacos , Animais , Bases de Dados Genéticas , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Terapia de Alvo Molecular , Invasividade Neoplásica , Células PC-3 , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Receptores Purinérgicos P2X4/genética , Receptores Purinérgicos P2X4/metabolismo , Transdução de Sinais , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Anticancer Drugs ; 33(3): 300-307, 2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-34924498

RESUMO

Small cell lung cancer (SCLC) is frequently found disseminated at first presentation and holds a poor prognosis due to emerging resistance to first-line platinum-based and second-line topotecan chemotherapy. The present investigation tested the antitumor activity of rovalpituzumab tesirine (ROVA-T), a cytotoxic anti-DLL3 drug conjugate, against two SCLC and a corresponding SCLC CTC cell line established from a ROVA-T-resistant patient to characterize the mechanism of recurrence. Two cell lines were established from an SCLC patient progressing under ROVA-T therapy and characterized with respect to chemosensitivity against this drug as well as against currently applied chemotherapeutics and for their delta-like 3 (DLL3) expression. The chemosensitivity assays demonstrate that most SCLC lines show IC50 values exceeding the ROVA-T in-vivo concentrations and that slow-growing cells and lines showing spheroidal growth or proliferation as corresponding circulating tumor cells (CTCs) exhibit higher resistance. Chemosensitivity of the cell lines is not correlated with DLL3 protein expression possibly due to toxicity of the free payload in tissue culture. The clinical trials and experimental results demonstrate that refractoriness to ROVA-T is linked to a low initial tumor expression of DLL3, loss of DLL3 expression, higher chemoresistance to ROVA-T and the putative formation of resistant spheroids by the SCLC cells.


Assuntos
Antineoplásicos , Neoplasias Pulmonares , Células Neoplásicas Circulantes , Carcinoma de Pequenas Células do Pulmão , Anticorpos Monoclonais Humanizados , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Benzodiazepinonas/uso terapêutico , Linhagem Celular Tumoral , Humanos , Imunoconjugados , Peptídeos e Proteínas de Sinalização Intracelular/uso terapêutico , Neoplasias Pulmonares/patologia , Proteínas de Membrana/metabolismo , Carcinoma de Pequenas Células do Pulmão/patologia
5.
Chem Pharm Bull (Tokyo) ; 70(2): 106-110, 2022 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-34897163

RESUMO

Benzolactams have unique biological activity and high utility in the synthesis of valuable compounds with direct applicability to oxindole alkaloids and antibacterial agents. Despite recent advances in organic chemistry and the growing number of reported methods for synthesizing benzolactams, their preparation still requires a multistep process. C-H amination reactions can convert aromatic C(sp2)-H bonds directly to C(sp2)-N bonds, and this direct approach to C-N bond formation offers effective access to benzolactams. Hypervalent iodine reagents are promising tools for achieving oxidative C-H amination. Motivated by our ongoing research efforts toward the development of useful hypervalent-iodine-mediated oxidative transformations, we herein describe an effective intramolecular oxidative C-H amination reaction based on µ-oxo hypervalent iodine catalysis for the synthesis of benzolactams bearing various functional groups.


Assuntos
Benzodiazepinonas/química , Carbono/química , Hidrogênio/química , Iodo/química , Aminação , Benzodiazepinonas/síntese química , Catálise , Ciclização , Oxirredução
6.
J Med Chem ; 64(24): 17901-17919, 2021 12 23.
Artigo em Inglês | MEDLINE | ID: mdl-34845907

RESUMO

Due to the neuroprotective role of the Na+/Ca2+ exchanger (NCX) isoforms NCX1 and NCX3, we synthesized novel benzodiazepinone derivatives of the unique NCX activator Neurounina-1, named compounds 1-19. The derivatives are characterized by a benzodiazepinonic nucleus linked to five- or six-membered cyclic amines via a methylene, ethylene, or acetyl spacer. The compounds have been screened on NCX1/NCX3 isoform activities by a high-throughput screening approach, and the most promising were characterized by patch-clamp electrophysiology and Fura-2AM video imaging. We identified two novel modulators of NCX: compound 4, inhibiting NCX1 reverse mode, and compound 14, enhancing NCX1 and NCX3 activity. Compound 1 displayed neuroprotection in two preclinical models of brain ischemia. The analysis of the conformational and steric features led to the identification of the molecular volume required for selective NCX1 activation for mixed NCX1/NCX3 activation or for NCX1 inhibition, providing the first prototypal model for the design of optimized isoform modulators.


Assuntos
Benzodiazepinonas/farmacologia , Fármacos Neuroprotetores/farmacologia , Isoformas de Proteínas/antagonistas & inibidores , Pirrolidinas/química , Trocador de Sódio e Cálcio/antagonistas & inibidores , Animais , Benzodiazepinonas/química , Desenho de Fármacos , Isoformas de Proteínas/metabolismo , Trocador de Sódio e Cálcio/metabolismo , Relação Estrutura-Atividade
7.
Biomolecules ; 11(9)2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34572579

RESUMO

SARS-CoV-2 virus mutations might increase its virulence, and thus the severity and duration of the ongoing pandemic. Global drug discovery campaigns have successfully developed several vaccines to reduce the number of infections by the virus. However, finding a small molecule pharmaceutical that is effective in inhibiting SARS-CoV-2 remains a challenge. Natural products are the origin of many currently used pharmaceuticals and, for this reason, a library of in-house fungal extracts were screened to assess their potential to inhibit the main viral protease Mpro in vitro. The extract of Penicillium citrinum, TDPEF34, showed potential inhibition and was further analysed to identify potential Mpro inhibitors. Following bio-guided isolation, a series of benzodiazepine alkaloids cyclopenins with good-to-moderate activity against SARS-CoV-2 Mpro were identified. The mode of enzyme inhibition of these compounds was predicted by docking and molecular dynamic simulation. Compounds 1 (isolated as two conformers of S- and R-isomers), 2, and 4 were found to have promising in vitro inhibitory activity towards Mpro, with an IC50 values range of 0.36-0.89 µM comparable to the positive control GC376. The in silico investigation revealed compounds to achieve stable binding with the enzyme active site through multiple H-bonding and hydrophobic interactions. Additionally, the isolated compounds showed very good drug-likeness and ADMET properties. Our findings could be utilized in further in vitro and in vivo investigations to produce anti-SARS-CoV-2 drug candidates. These findings also provide critical structural information that could be used in the future for designing potent Mpro inhibitors.


Assuntos
Proteases 3C de Coronavírus , Inibidores de Cisteína Proteinase , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Penicillium/química , SARS-CoV-2/enzimologia , Benzodiazepinonas/química , Benzodiazepinonas/isolamento & purificação , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/química , Inibidores de Cisteína Proteinase/química , Inibidores de Cisteína Proteinase/isolamento & purificação
9.
Nat Chem Biol ; 17(12): 1238-1244, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34556862

RESUMO

Cholecystokinin A receptor (CCKAR) belongs to family A G-protein-coupled receptors and regulates nutrient homeostasis upon stimulation by cholecystokinin (CCK). It is an attractive drug target for gastrointestinal and metabolic diseases. One distinguishing feature of CCKAR is its ability to interact with a sulfated ligand and to couple with divergent G-protein subtypes, including Gs, Gi and Gq. However, the basis for G-protein coupling promiscuity and ligand recognition by CCKAR remains unknown. Here, we present three cryo-electron microscopy structures of sulfated CCK-8-activated CCKAR in complex with Gs, Gi and Gq heterotrimers, respectively. CCKAR presents a similar conformation in the three structures, whereas conformational differences in the 'wavy hook' of the Gα subunits and ICL3 of the receptor serve as determinants in G-protein coupling selectivity. Our findings provide a framework for understanding G-protein coupling promiscuity by CCKAR and uncover the mechanism of receptor recognition by sulfated CCK-8.


Assuntos
Colecistocinina/química , Receptor de Colecistocinina A/química , Receptores Acoplados a Proteínas G/química , Sincalida/análogos & derivados , Sequência de Aminoácidos , Benzodiazepinonas/química , Microscopia Crioeletrônica , Humanos , Ligantes , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Multimerização Proteica , Sincalida/química , Triazóis/química
10.
Front Cell Infect Microbiol ; 11: 686035, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34350133

RESUMO

The failure of highly active antiretroviral therapy (HAART) has been largely responsible for the existence of latent human immunodeficiency virus type 1 (HIV-1) reservoirs. The "shock and kill" strategy was confirmed to reactivate HIV-1 latent reservoirs by latency-reversing agents (LRAs) for accelerated HIV-1 clearance. However, a single LRA might be insufficient to induce HIV-1 reactivation from latency due to the complexity of the multiple signaling regulatory pathways that establish the HIV-1 latent reservoir. Therefore, combinations of LRAs or dual-mechanism LRAs are urgently needed to purge the latent reservoirs. We demonstrate here for the first time that a dual-target inhibitor with a specific suppressive effect on both BRD4 and TIP60, CPI-637, could reactivate latent HIV-1 in vitro by permitting Tat to bind positive transcription elongation factor b (P-TEFb) and assembling Tat-super-elongation complex (SEC) formation. In addition, CPI-637-mediated TIP60 downregulation further stimulated BRD4 dissociation from the HIV-1 long terminal repeat (LTR) promoter, allowing Tat to more effectively bind P-TEFb compared to BRD4 inhibition alone. Much more importantly, CPI-637 exerted a potent synergistic effect but alleviated global T cell activation and blocked viral spread to uninfected bystander CD4+ T cells with minimal cytotoxicity. Our results indicate that CPI-637 opens up the prospect of novel dual-target inhibitors for antagonizing HIV-1 latency and deserves further investigation for development as a promising LRA with a "shock and kill" strategy.


Assuntos
Benzodiazepinonas/farmacologia , Infecções por HIV , HIV-1 , Linfócitos T CD4-Positivos , Proteínas de Ciclo Celular/antagonistas & inibidores , Infecções por HIV/tratamento farmacológico , Humanos , Lisina Acetiltransferase 5/antagonistas & inibidores , Proteínas Nucleares , Fatores de Transcrição/antagonistas & inibidores , Ativação Viral , Latência Viral
11.
J Neuroinflammation ; 18(1): 184, 2021 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-34425835

RESUMO

BACKGROUND: White matter injury (WMI) is a major neuropathological event associated with intracerebral hemorrhage (ICH). P2X purinoreceptor 4 (P2X4R) is a member of the P2X purine receptor family, which plays a crucial role in regulating WMI and neuroinflammation in central nervous system (CNS) diseases. Our study investigated the role of P2X4R in the WMI and the inflammatory response in mice, as well as the possible mechanism of action after ICH. METHODS: ICH was induced in mice via collagenase injection. Mice were treated with 5-BDBD and ANA-12 to inhibit P2X4R and tropomyosin-related kinase receptor B (TrkB), respectively. Immunostaining and quantitative polymerase chain reaction (qPCR) were performed to detect microglial phenotypes after the inhibition of P2X4R. Western blots (WB) and immunostaining were used to examine WMI and the underlying molecular mechanisms. Cylinder, corner turn, wire hanging, and forelimb placement tests were conducted to evaluate neurobehavioral function. RESULTS: After ICH, the protein levels of P2X4R were upregulated, especially on day 7 after ICH, and were mainly located in the microglia. The inhibition of P2X4R via 5-BDBD promoted neurofunctional recovery after ICH as well as the transformation of the pro-inflammatory microglia induced by ICH into an anti-inflammatory phenotype, and attenuated ICH-induced WMI. Furthermore, we found that TrkB blockage can reverse the protective effects of WMI as well as neuroprotection after 5-BDBD treatment. This result indicates that P2X4R plays a crucial role in regulating WMI and neuroinflammation and that P2X4R inhibition may benefit patients with ICH. CONCLUSIONS: Our results demonstrated that P2X4R contributes to WMI by polarizing microglia into a pro-inflammatory phenotype after ICH. Furthermore, the inhibition of P2X4R promoted pro-inflammatory microglia polarization into an anti-inflammatory phenotype, enhanced brain-derived neurotrophic factor (BDNF) production, and through the BDNF/TrkB pathway, attenuated WMI and improved neurological function. Therefore, the regulation of P2X4R activation may be beneficial for the reducing of ICH-induced brain injury.


Assuntos
Hemorragia Cerebral/patologia , Microglia/efeitos dos fármacos , Atividade Motora/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Antagonistas do Receptor Purinérgico P2X/farmacologia , Substância Branca/efeitos dos fármacos , Animais , Benzodiazepinonas/farmacologia , Hemorragia Cerebral/metabolismo , Modelos Animais de Doenças , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Microglia/metabolismo , Microglia/patologia , Proteínas Tirosina Quinases/metabolismo , Substância Branca/metabolismo , Substância Branca/patologia
13.
Neurobiol Dis ; 159: 105480, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34411705

RESUMO

Imbalance in cellular ionic homeostasis is a hallmark of several neurodegenerative diseases including Amyotrophic Lateral Sclerosis (ALS). Sodium-calcium exchanger (NCX) is a membrane antiporter that, operating in a bidirectional way, couples the exchange of Ca2+ and Na + ions in neurons and glial cells, thus controlling the intracellular homeostasis of these ions. Among the three NCX genes, NCX1 and NCX2 are widely expressed within the CNS, while NCX3 is present only in skeletal muscles and at lower levels of expression in selected brain regions. ALS mice showed a reduction in the expression and activity of NCX1 and NCX2 consistent with disease progression, therefore we aimed to investigate their role in ALS pathophysiology. Notably, we demonstrated that the pharmacological activation of NCX1 and NCX2 by the prolonged treatment of SOD1G93A mice with the newly synthesized compound neurounina: (1) prevented the reduction in NCX activity observed in spinal cord; (2) preserved motor neurons survival in the ventral spinal horn of SOD1G93A mice; (3) prevented the spinal cord accumulation of misfolded SOD1; (4) reduced astroglia and microglia activation and spared the resident microglia cells in the spinal cord; (5) improved the lifespan and mitigated motor symptoms of ALS mice. The present study highlights the significant role of NCX1 and NCX2 in the pathophysiology of this neurodegenerative disorder and paves the way for the design of a new pharmacological approach for ALS.


Assuntos
Esclerose Amiotrófica Lateral/metabolismo , Benzodiazepinonas/farmacologia , Neurônios Motores/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Pirrolidinas/farmacologia , Trocador de Sódio e Cálcio/agonistas , Medula Espinal/efeitos dos fármacos , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/patologia , Esclerose Amiotrófica Lateral/fisiopatologia , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Astrócitos/patologia , Humanos , Camundongos , Camundongos Transgênicos , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/patologia , Neurônios Motores/metabolismo , Neurônios Motores/patologia , /fisiopatologia , Medula Espinal/metabolismo , Medula Espinal/patologia , Superóxido Dismutase/genética , Taxa de Sobrevida
14.
Neurol Res ; 43(12): 1107-1115, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34461817

RESUMO

OBJECTIVE: To investigate the histopathological effects of a peripheral benzodiazepine receptor agonist (Ro5-4864) on epidural fibrosis (EF) in an experimental study model (post-laminectomy) in rats. METHODS: A total of 32 albino Wistar rats were randomly divided into four equal groups (n = 8). In Group 1, no treatment was applied after laminectomy (control group). In Group 2, hemostasis was achieved after Laminectomy, and the surgical procedure was terminated by placing a 2-mm absorbable gelatin sponge dipped in saline into the epidural space. In Group 3, low-dose (4 mg/kg) Ro5-4864 was administered 30 minutes before the surgery. In Group 4, high-dose (8 mg/kg) Ro5-4864 was administered 30 minutes before the surgery. A histopathological examination was performed to evaluate arachnoidal invasion and EF. RESULTS: Our data revealed the EF was significantly reduced in rats treated with high-dose Ro5-4864 (Group 4) compared to the control and saline-soaked Spongostan groups (p = 0.000 and p = 0.006, respectively). There was no significant difference between the groups treated with high- and low-dose Ro5-4864. Arachnoidal invasion was not seen in any of the rats in the high-dose R05-4864 group. However, the arachnoidal invasion results did not significantly differ between the study groups (p = 0.052 = 0.05). CONCLUSIONS: Our study showed that Ro5-4864 could be effective in reducing EF in rats after.


Assuntos
Benzodiazepinonas/farmacologia , Espaço Epidural/patologia , Laminectomia/efeitos adversos , Aderências Teciduais/prevenção & controle , Animais , Modelos Animais de Doenças , Síndrome Pós-Laminectomia/prevenção & controle , Feminino , Ratos , Ratos Wistar
15.
Cancer Treat Res Commun ; 28: 100405, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34329846

RESUMO

BACKGROUND: Agents targeting programmed cell death protein 1 (PD-1) have been approved as monotherapy for patients with small cell lung cancer (SCLC). In preclinical models, the combined targeting of PD-1 and delta-like protein 3 resulted in enhanced antitumor activity. Herein, we report results from the expansion arm of study NCT03000257 evaluating the combination of the anti-PD-1 antibody budigalimab and the targeted antibody-drug conjugate rovalpituzumab tesirine (Rova-T) in patients with previously treated SCLC. MATERIALS AND METHODS: This expansion arm of a multicenter, open-label, multi-arm, first-in-human phase 1 clinical trial enrolled adult patients with progressive SCLC. The primary objective was to assess safety and tolerability. Patients received budigalimab 375 mg via intravenous infusion every 3 weeks, and Rova-T was administered as a dose of 0.3 mg/kg intravenously, on day 1 of the first and third 3-week cycle. RESULTS: As of October 2019, 31 patients with SCLC were enrolled and treated with budigalimab plus Rova-T. The combination was tolerated, with the most common treatment-emergent adverse events (in >30%) being pleural effusion, fatigue, and cough. The overall response rate was 24.1%, with one confirmed complete response and six confirmed partial responses. The overall response rate in patients with high delta-like protein 3 expression was similar (21.1%). The median progression-free survival was 3.48 months. CONCLUSION: Combination therapy with budigalimab and Rova-T had promising efficacy and appeared to be tolerated in patients with SCLC. Although Rova-T development has been discontinued, development of budigalimab combined with other anticancer agents is ongoing. CLINICAL TRIAL REGISTRATION NUMBER: NCT03000257 Statement on originality of the work The manuscript represents original work and has not been submitted for publication elsewhere nor previously published. Statement of prior presentation Data from this study were previously presented at the European Society for Medical Oncology (ESMO) Congress 2019.


Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Benzodiazepinonas/uso terapêutico , Imunoconjugados/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Adulto , Idoso , Anticorpos Monoclonais Humanizados/farmacocinética , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Benzodiazepinonas/farmacocinética , Feminino , Humanos , Imunoconjugados/farmacocinética , Masculino , Pessoa de Meia-Idade
16.
J Thorac Oncol ; 16(9): 1582-1588, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34242790

RESUMO

INTRODUCTION: Rovalpituzumab tesirine (Rova-T) is an antibody-drug conjugate targeting DLL3, a Notch pathway ligand highly expressed on SCLC cells. Rova-T was evaluated alone or in combination with platinum-based chemotherapy (cisplatin or carboplatin combined with etoposide [CE]) in frontline treatment of extensive-stage SCLC. METHODS: One cycle of CE pre-enrollment was permitted (later mandated). The following four cohorts were enrolled: Rova-T monotherapy (0.3 mg/kg, every 6 [q6] wk × 2; cohort 1; n = 4); Rova-T induction (0.3 mg/kg, q6 wk × 2) followed by CE every 21 days (q21) × 4 (cohort 2; n = 5); Rova-T (0.1 or 0.2 mg/kg, q6 wk × 2) overlapping with CE q21 × 4 (cohort 3; n = 14); and Rova-T maintenance (0.3 mg/kg, q6 wk × 2) after CE q21 × 4 (cohort 4; n = 3). RESULTS: A total of 26 patients were dosed (cohort 3: 14; cohorts 1, 2, and 4 combined: 12). Median age was 66 years, and 73% had Eastern Cooperative Oncology Group performance status of 1. In cohort 3, seven patients (50%) had confirmed objective responses, with a median progression-free survival of 5.2 months and median overall survival of 10.3 months. Compared with cohorts 1, 2, and 4 combined, cohort 3 had lower frequency of some Rova-T-related adverse events of special interest, such as pleural effusion (0 versus 33%), pericardial effusion (0 versus 17%), ascites (0 versus 8%), peripheral edema (36% versus 42%), generalized edema (0 versus 8%), pneumonia (7% versus 25%), and hypoalbuminemia (0 versus 17%). CONCLUSIONS: Lower Rova-T doses may be associated with lower incidence of some Rova-T-related adverse events of special interest. Rova-T 0.2 mg/kg plus CE (cohort 3) was tolerable; however, there was no clear efficacy benefit of adding Rova-T to CE.


Assuntos
Imunoconjugados , Neoplasias Pulmonares , Carcinoma de Pequenas Células do Pulmão , Idoso , Anticorpos Monoclonais Humanizados/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Benzodiazepinonas/uso terapêutico , Carboplatina/uso terapêutico , Etoposídeo/uso terapêutico , Humanos , Imunoconjugados/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico
17.
J Med Chem ; 64(14): 10102-10123, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34255515

RESUMO

CREBBP (CBP/KAT3A) and its paralogue EP300 (KAT3B) are lysine acetyltransferases (KATs) that are essential for human development. They each comprise 10 domains through which they interact with >400 proteins, making them important transcriptional co-activators and key nodes in the human protein-protein interactome. The bromodomains of CREBBP and EP300 enable the binding of acetylated lysine residues from histones and a number of other important proteins, including p53, p73, E2F, and GATA1. Here, we report a work to develop a high-affinity, small-molecule ligand for the CREBBP and EP300 bromodomains [(-)-OXFBD05] that shows >100-fold selectivity over a representative member of the BET bromodomains, BRD4(1). Cellular studies using this ligand demonstrate that the inhibition of the CREBBP/EP300 bromodomain in HCT116 colon cancer cells results in lowered levels of c-Myc and a reduction in H3K18 and H3K27 acetylation. In hypoxia (<0.1% O2), the inhibition of the CREBBP/EP300 bromodomain results in the enhanced stabilization of HIF-1α.


Assuntos
Benzodiazepinonas/farmacologia , Proteína de Ligação a CREB/antagonistas & inibidores , Desenho de Fármacos , Proteína p300 Associada a E1A/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/farmacologia , Benzodiazepinonas/síntese química , Benzodiazepinonas/química , Proteína de Ligação a CREB/metabolismo , Relação Dose-Resposta a Droga , Proteína p300 Associada a E1A/metabolismo , Células HCT116 , Humanos , Ligantes , Estrutura Molecular , Bibliotecas de Moléculas Pequenas/síntese química , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade
18.
Cancer Sci ; 112(8): 2984-2992, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34107132

RESUMO

Delta-like canonical Notch ligand 3 (DLL3) is a member of the Delta/Serrate/Lag2 (DSL) Notch receptor ligand family and plays a crucial role in Notch signaling, which influences various cellular processes including differentiation, proliferation, survival, and apoptosis. DLL3 is expressed throughout the presomitic mesoderm and is localized to the rostral somatic compartments; mutations in DLL3 induce skeletal abnormalities such as spondylocostal dysostosis. Recently, DLL3 has attracted interest as a novel molecular target due to its high expression in neuroendocrine carcinoma of the lung. Moreover, a DLL3-targeting Ab-drug conjugate, rovalpituzumab tesirine (ROVA-T), has been developed as a new treatment with proven antitumor activity. However, the development of ROVA-T was suspended because of shorter overall survival compared to topotecan, the second-line standard treatment. Thus, several studies on the mechanism and function of DLL3 in several malignancies are underway to find a new strategy for targeting DLL3. In this review, we discuss the roles of DLL3 in various malignancies and the future perspectives of DLL3-related research, especially as a therapeutic target.


Assuntos
Antineoplásicos/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Membrana/genética , Neoplasias/tratamento farmacológico , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos/uso terapêutico , Benzodiazepinonas/farmacologia , Benzodiazepinonas/uso terapêutico , Ensaios Clínicos como Assunto , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Peptídeos e Proteínas de Sinalização Intracelular/efeitos dos fármacos , Masculino , Proteínas de Membrana/efeitos dos fármacos , Terapia de Alvo Molecular , Mutação , Neoplasias/genética
19.
Bioorg Chem ; 114: 105081, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34153811

RESUMO

Selective inhibition of histone deacetylase 6 (HDAC6) has been emerged as a promising approach to cancer treatment. As a pivotal strategy for drug discovery,molecular hybridization was introduced in this study and a series of pyrrolo[2,1-c][1,4] benzodiazepine-3,11-diones (PBDs) based hydroxamic acids was rationally designed and synthesizedas novel selective HDAC6 inhibitors. Preliminary in vitro enzyme inhibition assay and structure-activity relationship (SAR) discussion confirmed our design strategy and met the expectation. Several of the compounds showed high potent against HDAC6 enzyme in vitro, and compound A7 with a long aliphatic linker was revealed to have the similar activity as the positive control tubastatin A. Further in vitro characterization of A7 demonstrates the metastasis inhibitory potency in MDA-MB-231 cell line and western blotting showed that A7 could induce the upregulation of Ac-α-tubulin, but not induce the excessive acetylation of histone H3, which indicated that the compound had HDAC6 targeting effect in MDA-MB-231 cells. In vivo study revealed that compound A7 has satisfactory inhibitory effects onliver and lung metastasis of breast cancer in mice. Molecular docking released that A7 could fit well with the receptor and interact with some key residues, which lays a foundation for further structural modifications to elucidate the interaction mode between compounds and target protein. This pharmacological investigation workflow provided a reasonable and reference methodto examine the pharmacological effects of inhibiting HDAC6 with a single molecule, either in vitro or in vivo. All of these results suggested that A7 is a promising lead compound that could lead to the further development of novel selective HDAC6 inhibitors for the treatment of tumor metastasis.


Assuntos
Antineoplásicos/farmacologia , Benzodiazepinonas/farmacologia , Desacetilase 6 de Histona/antagonistas & inibidores , Inibidores de Histona Desacetilases/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Benzodiazepinonas/síntese química , Benzodiazepinonas/química , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Desacetilase 6 de Histona/metabolismo , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/química , Humanos , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais Cultivadas
20.
Purinergic Signal ; 17(3): 425-438, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33966147

RESUMO

Ischemic stroke is the most serious disease that harms human beings. In principle, its treatment is to restore blood flow supply as soon as possible. However, after the blood flow is restored, it will lead to secondary brain injury, that is, ischemia-reperfusion injury. The mechanism of ischemia-reperfusion injury is very complicated. This study showed that P2X4 receptors in the pyramidal neurons of rat hippocampus were significantly upregulated in the early stage of ischemia-reperfusion injury. Neurons with high expression of P2X4 receptors are neurons that are undergoing apoptosis. Intraventricular injection of the P2X4 receptor antagonist 5-(3-bromophenyl)-1,3-dihydro-2H-benzofuro[3,2-e]-1,4-diazepin-2-one (5-BDBD) and PSB-12062 can partially block neuronal apoptosis, to promote the survival of neurons, indicating that ATP through P2X4 receptors is involved in the process of cerebral ischemia-reperfusion injury. Therefore, identifying the mechanism of neuronal degeneration induced by extracellular ATP via P2X4 receptors after ischemia-reperfusion will likely find new targets for the treatment of ischemia-reperfusion injury, and will provide a useful theoretical basis for the treatment of ischemia-reperfusion injury.


Assuntos
Isquemia Encefálica/metabolismo , Região CA1 Hipocampal/metabolismo , Células Piramidais/metabolismo , Receptores Purinérgicos P2X4/biossíntese , Traumatismo por Reperfusão/metabolismo , Animais , Benzodiazepinonas/farmacologia , Isquemia Encefálica/genética , Isquemia Encefálica/patologia , Região CA1 Hipocampal/efeitos dos fármacos , Expressão Gênica , Masculino , Antagonistas do Receptor Purinérgico P2X/farmacologia , Células Piramidais/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores Purinérgicos P2X4/genética , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/patologia
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