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1.
Nature ; 613(7944): 582-587, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36599980

RESUMO

Cas12a2 is a CRISPR-associated nuclease that performs RNA-guided, sequence-nonspecific degradation of single-stranded RNA, single-stranded DNA and double-stranded DNA following recognition of a complementary RNA target, culminating in abortive infection1. Here we report structures of Cas12a2 in binary, ternary and quaternary complexes to reveal a complete activation pathway. Our structures reveal that Cas12a2 is autoinhibited until binding a cognate RNA target, which exposes the RuvC active site within a large, positively charged cleft. Double-stranded DNA substrates are captured through duplex distortion and local melting, stabilized by pairs of 'aromatic clamp' residues that are crucial for double-stranded DNA degradation and in vivo immune system function. Our work provides a structural basis for this mechanism of abortive infection to achieve population-level immunity, which can be leveraged to create rational mutants that degrade a spectrum of collateral substrates.


Assuntos
Proteínas Associadas a CRISPR , Sistemas CRISPR-Cas , RNA , Proteínas Associadas a CRISPR/antagonistas & inibidores , Proteínas Associadas a CRISPR/metabolismo , DNA/química , DNA/imunologia , DNA/metabolismo , RNA/química , RNA/metabolismo , Ativação Enzimática , Domínio Catalítico , Especificidade por Substrato
3.
BMC Psychiatry ; 23(1): 17, 2023 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-36624454

RESUMO

BACKGROUND: Maladaptation of the HPA (hypothalamic-pituitary-adrenal) axis plays an important role in depression-like behaviour, but the specific molecular mechanisms are unknown. Here, we determined the roles of CRHR1 (corticotrophin releasing hormone receptor 1) and nectin3 in LPS (lipopolysaccharide)-induced depression-like behaviour in mice. METHODS: C57BL/6 male mice were intraperitoneally injected with LPS (0.83 g/kg), and the open field, novelty-suppressed feeding, forced swimming, and tail suspension tests were performed after intraperitoneal injections of saline or antalarmin (20 mg/kg). The hippocampal mRNA levels of CRHR1 and nectin3 were determined by quantitative reverse transcription-PCR. The hippocampal protein levels of CRHR1, nectin3, and calbindin were measured by western blotting. The CORT (corticosterone) levels in the blood were measured by ELISA kits. RESULTS: Antalarmin alleviated LPS-induced depression-like behaviour in male mice. Furthermore, antalarmin significantly inhibited changes in CRHR1, nectin3 and calbindin levels in the hippocampus and reduced the increase in CORT levels in LPS-treated mice. CONCLUSION: CRHR1antagonist showed antidepressant effects in LPS-induced depressive mice, and CRHR1/nectin3 signalling may play a crucial role in this process.


Assuntos
Depressão , Receptores de Hormônio Liberador da Corticotropina , Animais , Masculino , Camundongos , Antidepressivos/farmacologia , Antidepressivos/uso terapêutico , Depressão/induzido quimicamente , Depressão/tratamento farmacológico , Modelos Animais de Doenças , Hipocampo , Sistema Hipotálamo-Hipofisário/metabolismo , Lipopolissacarídeos , Camundongos Endogâmicos C57BL , Sistema Hipófise-Suprarrenal/metabolismo , Receptores de Hormônio Liberador da Corticotropina/antagonistas & inibidores
4.
J Enzyme Inhib Med Chem ; 38(1): 2163242, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36629431

RESUMO

Androgen deprivation therapy (ADT) is a common treatment for recurrent prostate cancer (PC). However, after a certain period of responsiveness, ADT resistance occurs virtually in all patients and the disease progresses to lethal metastatic castration-resistant prostate cancer (mCRPC). Aberrant expression and function of the epigenetic modifiers EZH2 and BET over activates c-myc, an oncogenic transcription factor critically contributing to mCRPC. In the present work, we tested, for the first time, the combination of an EZH2 inhibitor with a BET inhibitor in metastatic PC cells. The combination outperformed single drugs in inhibiting cell viability, cell proliferation and clonogenic ability, and concomitantly reduced both c-myc and NF-kB expression. Although these promising results will warrant further in vivo validation, they represent the first step to establishing the rationale that the proposed combination might be suitable for mCRPC treatment, by exploiting molecular targets different from androgen receptor.


Assuntos
Antineoplásicos , Neoplasias de Próstata Resistentes à Castração , Humanos , Masculino , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proliferação de Células , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Fatores de Transcrição , Betaína-Homocisteína S-Metiltransferase/antagonistas & inibidores , Betaína-Homocisteína S-Metiltransferase/metabolismo
5.
Sci Rep ; 13(1): 1378, 2023 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-36697438

RESUMO

Targeted therapy significantly impairs tumour growth but suffers from limitations, among which the 'flare' ('rebound') effect. Among cancers driven by tyrosine kinase receptors, those relying on alterations of the MET oncogene benefit from treatment by specific inhibitors. Previously, we reported that discontinuation of MET tyrosine kinase receptor inhibition causes 'rebound' activation of the oncogene, with a post-treatment transient hyperphosphorylation phase that culminates into a dramatic increase in cancer cell proliferation. The molecular mechanisms behind the 'MET burst' after treatment cessation are unknown but critically important for patients. Here we identify a positive feedback loop mediated by the AKT/mTOR pathway leading to (a) enhanced MET translation by activating p70S6K and 4EBP1 and (b) MET hyper-phosphorylation by inactivation of the tyrosine-phosphatase PTP1B. The latter effect is due to m-TOR-driven PTP1B phosphorylation of the inhibitory residues Ser50 and Ser378. These data provide in vitro evidence for the use of mTOR inhibitors to prevent the 'flare effect' in MET targeted therapy, with potential applicative ramifications for patient clinical management.


Assuntos
Neoplasias , Proteínas Proto-Oncogênicas c-akt , Proteínas Proto-Oncogênicas c-met , Serina-Treonina Quinases TOR , Humanos , Linhagem Celular Tumoral , Neoplasias/tratamento farmacológico , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo , Retroalimentação Fisiológica
6.
J Pharm Biomed Anal ; 225: 115211, 2023 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-36603395

RESUMO

The cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors palbociclib, ribociclib, and abemaciclib were approved by the U.S. Food and Drug Administration (FDA) and European Medicine Agency for the treatment of breast cancer between 2015 and 2018. Oral tumor therapeutics extend the options for cancer therapy, but also challenge physicians and patients. The aim of the present work was to establish a semi-automated liquid-chromatography tandem mass spectrometry (LC-MS/MS) method for the simultaneous quantification of abemaciclib, its active metabolites abemaciclib M20 and M2, palbociclib, and ribociclib in human serum. Detuning of ribociclib enabled the development of a simultaneous quantification method for abemaciclib, M20, M2, palbociclib, and ribociclib in the respective relevant concentration ranges based on semi-automated sample preparation with isotope dilution LC-MS/MS. The method was validated according to the guidance of the FDA. The LC-MS/MS method was successfully validated according to FDA and showed inaccuracies ≤ 10.7% and imprecisions ≤ 8.51%. Linearity was given from 20 to 800 ng/mL for abemaciclib, 15-600 ng/mL for M20, 10-400 ng/mL for M2 and palbociclib, and 100-4000 ng/mL for ribociclib. Normalized matrix factors and process efficiency showed no significant matrix effects regardless of the analytes. To demonstrate the applicability of the method, authentic samples were also analyzed. This novel semi-automated LC-MS/MS method covering all previously approved CDK4/6 inhibitors as well as the similarly pharmacologically active metabolites in human serum simultaneously was developed for potential future use in routine analysis in order to improve personalized therapy, patient safety, and treatment success.


Assuntos
Neoplasias da Mama , Feminino , Humanos , Aminopiridinas/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Cromatografia Líquida , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Monitoramento de Medicamentos , Inibidores de Proteínas Quinases , Espectrometria de Massas em Tandem/métodos , Quinase 6 Dependente de Ciclina/antagonistas & inibidores
7.
BMC Med ; 21(1): 2, 2023 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-36600247

RESUMO

BACKGROUND: HL-085 is a selective, orally administered MEK1/2 inhibitor. We aimed to evaluate the safety and efficacy of HL-085 in patients with advanced melanoma harboring NRAS mutations. METHODS: This was a multicenter phase 1 study. HL-085 was administered twice daily in a standard 3 + 3 dose-escalation design (10 dose cohorts; 0.5-18 mg twice daily), followed by dose expansion at the recommended phase II dose (RP2D). The primary endpoints included tolerability, dose-limiting toxicity (DLT), maximum tolerated dose (MTD) and RP2D. RESULTS: Between September 13, 2017, and January 18, 2021, 42 patients were enrolled (dose escalation phase: n = 30; dose expansion phase: n = 12). No DLT was reported during dose escalation and MTD was not reached with HL-085 doses up to 18 mg twice daily. The RP2D was 12 mg twice daily. The most common all-grade drug-related adverse events (AEs) across all dose levels were rash (61.9%), increased creatine phosphokinase (CK, 59.5%), face edema (50.0%), increased aspartate aminotransferase (47.6%), peripheral edema (40.5%), diarrhea (33.3%), alanine aminotransferase (33.3%), and paronychia (19.0%), most of which were grade 1 and 2. Most frequency of grade ≥ 3 AEs were CK (14.2%), asthenia (7.1%), peripheral edema (4.8%), and acneiform dermatitis (4.8%). In the cohort of 12 mg twice daily dose (15 patients), confirmed objective response rate was 26.7%; disease control rate was 86.7%; median duration of response was 2.9 months; median progression-free survival was 3.6 months. CONCLUSIONS: The HL-085 showed acceptable tolerability and substantial clinical activity in patients with advanced melanoma harboring NRAS mutations. TRIAL REGISTRATION: Trial registration ClinicalTrials.gov number: NCT03973151.


Assuntos
Melanoma , Quinases de Proteína Quinase Ativadas por Mitógeno , Humanos , Antineoplásicos/efeitos adversos , Antineoplásicos/uso terapêutico , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/uso terapêutico , Melanoma/tratamento farmacológico , Melanoma/genética , Proteínas de Membrana/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Mutação , Intervalo Livre de Progressão , Inibidores de Proteínas Quinases/efeitos adversos , Inibidores de Proteínas Quinases/uso terapêutico
8.
J Neuroinflammation ; 20(1): 4, 2023 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-36600259

RESUMO

BACKGROUND: Ischemic stroke immediately evokes a strong neuro-inflammatory response within the vascular compartment, which contributes to primary infarct development under vessel occlusion as well as further infarct growth despite recanalization, referred to as ischemia/reperfusion injury. Later, in the subacute phase of stroke (beyond day 1 after recanalization), further inflammatory processes within the brain parenchyma follow. Whether this second wave of parenchymal inflammation contributes to an additional/secondary increase in infarct volumes and bears the potential to be pharmacologically targeted remains elusive. We addressed the role of the NLR-family pyrin domain-containing protein 3 (NLRP3) inflammasome in the subacute phase of ischemic stroke. METHODS: Focal cerebral ischemia was induced in C57Bl/6 mice by a 30-min transient middle cerebral artery occlusion (tMCAO). Animals were treated with the NLRP3 inhibitor MCC950 therapeutically 24 h after or prophylactically before tMCAO. Stroke outcome, including infarct size and functional deficits as well as the local inflammatory response, was assessed on day 7 after tMCAO. RESULTS: Infarct sizes on day 7 after tMCAO decreased about 35% after delayed and about 60% after prophylactic NLRP3 inhibition compared to vehicle. Functionally, pharmacological inhibition of NLRP3 mitigated the local inflammatory response in the ischemic brain as indicated by reduction of infiltrating immune cells and reactive astrogliosis. CONCLUSIONS: Our results demonstrate that the NLRP3 inflammasome continues to drive neuroinflammation within the subacute stroke phase. NLRP3 inflammasome inhibition leads to a better long-term outcome-even when administered with a delay of 1 day after stroke induction, indicating ongoing inflammation-driven infarct progression. These findings may pave the way for eagerly awaited delayed treatment options in ischemic stroke.


Assuntos
Isquemia Encefálica , Inflamassomos , AVC Isquêmico , Traumatismo por Reperfusão , Acidente Vascular Cerebral , Animais , Camundongos , Isquemia Encefálica/metabolismo , Infarto da Artéria Cerebral Média/complicações , Infarto da Artéria Cerebral Média/tratamento farmacológico , Infarto da Artéria Cerebral Média/metabolismo , Inflamassomos/antagonistas & inibidores , Inflamassomos/metabolismo , Inflamação/complicações , AVC Isquêmico/tratamento farmacológico , AVC Isquêmico/complicações , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Traumatismo por Reperfusão/metabolismo , Acidente Vascular Cerebral/metabolismo
9.
PLoS One ; 18(1): e0280526, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36652434

RESUMO

AP endonuclease 1 (APE1) processes DNA lesions including apurinic/apyrimidinic sites and 3´-blocking groups, mediating base excision repair and single strand break repair. Much effort has focused on developing specific inhibitors of APE1, which could have important applications in basic research and potentially lead to clinical anticancer agents. We used structural, biophysical, and biochemical methods to characterize several reported inhibitors, including 7-nitroindole-2-carboxylic acid (CRT0044876), given its small size, reported potency, and widespread use for studying APE1. Intriguingly, NMR chemical shift perturbation (CSP) experiments show that CRT0044876 and three similar indole-2-carboxylic acids bind a pocket distal from the APE1 active site. A crystal structure confirms these findings and defines the pose for 5-nitroindole-2-carboxylic acid. However, dynamic light scattering experiments show the indole compounds form colloidal aggregates that could bind (sequester) APE1, causing nonspecific inhibition. Endonuclease assays show the compounds lack significant APE1 inhibition under conditions (detergent) that disrupt aggregation. Thus, binding of the indole-2-carboxylic acids at the remote pocket does not inhibit APE1 repair activity. Myricetin also forms aggregates and lacks APE1 inhibition under aggregate-disrupting conditions. Two other reported compounds (MLS000552981, MLS000419194) inhibit APE1 in vitro with low micromolar IC50 and do not appear to aggregate in this concentration range. However, NMR CSP experiments indicate the compounds do not bind specifically to apo- or Mg2+-bound APE1, pointing to a non-specific mode of inhibition, possibly DNA binding. Our results highlight methods for rigorous interrogation of putative APE1 inhibitors and should facilitate future efforts to discover compounds that specifically inhibit this important repair enzyme.


Assuntos
Antineoplásicos , DNA Liase (Sítios Apurínicos ou Apirimidínicos) , Humanos , Antineoplásicos/farmacologia , Reparo do DNA , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/antagonistas & inibidores , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/química , Indóis/farmacologia
10.
Zhonghua Yi Xue Za Zhi ; 103(4): 265-270, 2023 Jan 31.
Artigo em Chinês | MEDLINE | ID: mdl-36660787

RESUMO

Objective: To explore the relationship between expression of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) inflammasome and improvement of macular structure in patients with wet age-related macular degeneration (wAMD) after anti-vascular endothelial growth factor (VEGF) therapy. Methods: A before-after study was carried out. A total of 110 patients (110 eyes) with wAMD who were admitted to Department of Ophthalmology, People's Hospital Affiliated to Shandong First Medical University between August 2019 and December 2021 were enrolled, and all patients were given vitreous injection of anti-VEGF drug (ranibizumab or bevacizumab). The aqueous humor was collected to detect mRNA levels of NLRP3, cysteinyl aspartate specific protease-1 (Caspase-1), apoptosis-associated speck-like protein (ASC) and interleukin (IL) 1ß by fluorescence quantitative PCR. The levels of IL-1ß, IL-18, tumor necrosis factor α (TNF-α) and VEGF in aqueous humor were detected by enzyme-linked immunosorbent assay (ELISA). The correlation between the above indexes and central macular thickness (CMT) in wAMD patients was analyzed by multivariate linear regression analysis. Results: In the 110 wAMD patients, there were 68 males and 42 females, with a mean age of (68.7±7.6) years. Compared with those before treatment, mRNA levels of NLRP3 (1.65±0.27, 1.34±0.19 vs 1.97±0.23, both P<0.017), Caspase-1 (1.47±0.15, 1.29±0.17 vs 1.53±0.18, both P<0.017), ASC (1.33±0.14, 1.21±0.18 vs 1.47±0.12, both P<0.017) and IL-1ß (1.78±0.21, 1.46±0.17 vs 2.21±0.24, both P<0.017), and levels of IL-1ß [(26.9±5.7), (20.3±4.6) vs (33.6±8.3) ng/L, both P<0.017], IL-18 [(32.7±7.6), (23.3±6.9) vs (46.4±9.4) ng/L, both P<0.017], TNF-α [(39.4±6.6), (21.7±6.3) vs (52.9±9.1) ng/L, both P<0.017] and VEGF [(35.7±10.2), (23.4±6.7) vs (65.4±19.3) ng/L, both P<0.017] were decreased after the first and second injection. Moreover, the above-mentioned indexes after second injection were lower than those after the first injection (all P<0.017). The results of multivariate linear regression analysis showed that NLRP3 mRNA (the first injection: ß=53.750, P<0.001; the second injection: ß=94.648, P<0.001), IL-1ß (the first injection: ß=1.356, P=0.021; the second injection: ß=2.008, P=0.003), IL-18 (the first injection: ß=1.984, P<0.001; the second injection: ß=1.251, P=0.003) and VEGF (the first injection: ß=1.875, P<0.001; the second injection: ß=2.119, P<0.001) had linear relationships with CMT. Conclusion: The decrease of NLRP3 inflammasome and its products in aqueous humor may be related to the improvement of macular structure in wAMD patients after anti-VEGF therapy.


Assuntos
Inflamassomos , Degeneração Macular , Fator A de Crescimento do Endotélio Vascular , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Caspase 1/metabolismo , Inflamassomos/metabolismo , Interleucina-18 , Interleucina-1beta/metabolismo , Degeneração Macular/tratamento farmacológico , Degeneração Macular/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , RNA Mensageiro , Fator de Necrose Tumoral alfa/metabolismo , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores
11.
Cell Death Dis ; 14(1): 25, 2023 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-36639658

RESUMO

Hepatocellular carcinoma (HCC) with lung metastasis is associated with poor prognosis and poor therapeutic outcomes. Studies have demonstrated that stiffened stroma can promote metastasis in various tumors. However, how the lung mechanical microenvironment favors circulating tumor cells remains unclear in metastatic HCC. Here, we found that the expression of cell migration-inducing hyaluronan-binding protein (CEMIP) was closely associated with lung metastasis and can promote pre-metastatic niche formation by increasing lung matrix stiffness. Furthermore, upregulated serum CEMIP was indicative of lung fibrotic changes severity in patients with HCC lung metastasis. By directly targeting CEMIP, pirfenidone can inhibit CEMIP/TGF-ß1/Smad signaling pathway and reduce lung metastases stiffening, demonstrating promising antitumor activity. Pirfenidone in combination with sorafenib can more effectively suppress the incidence of lung metastasis compared with sorafenib alone. This study is the first attempt to modulate the mechanical microenvironment for HCC therapy and highlights CEMIP as a potential target for the prevention and treatment of HCC lung metastasis. CEMIP mediating an HCC-permissive microenvironment through controlling matrix stiffness. Meanwhile, Pirfenidone could reduce metastasis stiffness and increases the anti-angiogenic effect of Sorafenib by directly targeting CEMIP.


Assuntos
Carcinoma Hepatocelular , Hialuronoglucosaminidase , Neoplasias Hepáticas , Neoplasias Pulmonares , Sorafenibe , Humanos , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/patologia , Transdução de Sinais , Sorafenibe/farmacologia , Sorafenibe/uso terapêutico , Microambiente Tumoral , Hialuronoglucosaminidase/antagonistas & inibidores
12.
Eur J Med Res ; 28(1): 28, 2023 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-36642707

RESUMO

BACKGROUND: Progestins can suppress endogenous luteinising hormone (LH) secretion from the pituitary gland and have shown similar efficacy in terms of collecting competent oocytes and embryos; however, some inconsistencies have been proposed regarding the quality of embryos collected with the use of progestins. This study aimed to evaluate euploidy rates and pregnancy outcomes in preimplantation genetic testing for aneuploidy (PGT-A) cycles using the progestin-primed ovarian stimulation (PPOS) protocol versus the gonadotropin-releasing hormone (GnRH) agonist/antagonist protocol. METHODS: This retrospective cohort study included 608 PGT-A cycles: 146 women in the PPOS group, 160 women in the GnRH agonist group, and 302 women in the GnRH antagonist group. This study was performed at the in vitro fertilisation (IVF) centre of Shanghai First Maternity and Infant Hospital between January 2019 and December 2021. Additionally, 267 corresponding first frozen embryo transfer (FET) cycles were analysed to assess pregnancy outcomes. RESULTS: The euploid blastocyst rate per injected metaphase II(MII) oocytes (14.60% vs. 14.09% vs. 13.94%) was comparable among the three groups (p > 0.05). No significant differences were observed among the three groups regarding pregnancy outcomes, including biochemical pregnancy, clinical pregnancy, ongoing pregnancy, implantation, miscarriage, ectopic pregnancy, and live birth rates per transfer in the first FET cycles (p > 0.05). CONCLUSIONS: The PPOS protocol had no negative effect on euploid blastocyst formation, and the pregnancy outcomes in FET cycles using the PPOS protocol were similar to those of the GnRH agonist and antagonist protocols. Trial registration This trial was retrospectively registered.


Assuntos
Aneuploidia , Testes Genéticos , Progestinas , Feminino , Humanos , Gravidez , China , Hormônio Liberador de Gonadotropina/agonistas , Hormônio Liberador de Gonadotropina/antagonistas & inibidores , Estudos Retrospectivos , Esteroides , Resultado da Gravidez , Técnicas de Reprodução Assistida
13.
Molecules ; 28(2)2023 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-36677690

RESUMO

Estradiol (E2) plays an important role in the progression of diseases such as breast cancer and endometriosis. Inhibition of 17ß-hydroxysteroid dehydrogenase type 1 (17ß-HSD1), the enzyme that catalyzes the last step in the biosynthesis of the estrogenic hormone E2, therefore constitutes an interesting approach for the treatment of these two estrogen-dependent diseases. In order to obtain new inhibitors of 17ß-HSD1, the impact of a m-carbamoylphenyloxy group at position three of an estrane nucleus was evaluated by preparing three derivatives of estrone (E1) and E2 using a microwave-assisted synthesis of diaryl ethers. Their inhibitory activity was addressed on two cell lines (T-47D and Z-12) representative of breast cancer and endometriosis, respectively, but unlike T-47D cells, Z-12 cells were not found suitable for testing potential 17ß-HSD1 inhibitors. Thus, the addition of the m-carbamoylphenyl group at C3 of E1 (compound 5) did not increase the inhibition of E1 to E2 transformation by 17ß-HSD1 present in T-47D cells (IC50 = 0.31 and 0.21 µM for 5 and E1, respectively), and this negative effect was more obvious for E2 derivatives 6 and 10 (IC50 = 1.2 and 1.3 µM, respectively). Molecular docking allowed us to identify key interactions with 17ß-HSD1 and to highlight these new inhibitors' actions through an opposite orientation than natural enzyme substrate E1's classical one. Furthermore, molecular modeling experiments explain the better inhibitory activity of E1-ether derivative 5, as opposed to the E2-ether derivatives 6 and 10. Finally, when tested on T-47D and Z-12 cells, compounds 5, 6 and 10 did not stimulate the proliferation of these two estrogen-dependent cell lines. In fact, they reduced it.


Assuntos
17-Hidroxiesteroide Desidrogenases , Neoplasias da Mama , Endometriose , Inibidores Enzimáticos , Feminino , Humanos , 17-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , Linhagem Celular Tumoral , Inibidores Enzimáticos/química , Estradiol , Estrogênios , Estrona/farmacologia , Simulação de Acoplamento Molecular
14.
Molecules ; 28(2)2023 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-36677536

RESUMO

A new series of Schiff-benzimidazole hybrids 3a-o has been designed and synthesized. The structure of the target compounds was proved by different spectroscopic and elemental analysis tools. The target compounds were evaluated for their in vitro cytotoxic activity against 60 cancer cell lines according to NCI single- and five-dose protocols. Consequently, four compounds were further examined against the most sensitive lung cancer A549 and NCI-H460 cell lines. Compounds 3e and 3g were the most active, achieving 3.58 ± 0.53, 1.71 ± 0.17 and 1.88 ± 0.35, 0.85 ± 0.24 against A549 and NCI-H460 cell lines, respectively. Moreover, they showed remarkable inhibitory activity on the VEGFR-2 TK with 86.23 and 89.89%, respectively, as compared with Sorafenib (88.17%). Moreover, cell cycle analysis of NCI-H460 cells treated with 3e and 3g showed cellular cycle arrest at both G1 and S phases (supported by caspases-9 study) with significant pro-apoptotic activity, as indicated by annexin V-FITC staining. The binding interactions of these compounds were confirmed through molecular docking studies; the most active compounds displayed complete overlay with, and a similar binding mode and pose to, Sorafenib, a reference VEGFR-2 inhibitor.


Assuntos
Antineoplásicos , Receptor 2 de Fatores de Crescimento do Endotélio Vascular , Antineoplásicos/química , Apoptose , Benzimidazóis/química , Proliferação de Células , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Simulação de Acoplamento Molecular , Estrutura Molecular , Bases de Schiff/farmacologia , Sorafenibe/farmacologia , Relação Estrutura-Atividade , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
15.
Molecules ; 28(2)2023 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-36677750

RESUMO

One crucial strategy for the treatment of breast cancer involves focusing on the Vascular Endothelial Growth Factor Receptor (VEGFR-2) signaling system. Consequently, the development of new (VEGFR-2) inhibitors is of the utmost importance. In this study, novel 3-thiazolhydrazinylcoumarins were designed and synthesized via the reaction of phenylazoacetylcoumarin with various hydrazonoyl halides and α-bromoketones. By using elemental and spectral analysis data (IR, 1H-NMR, 13C-NMR, and Mass), the ascribed structures for all newly synthesized compounds were clarified, and the mechanisms underlying their formation were delineated. The molecular docking studies of the resulting 6-(phenyldiazenyl)-2H-chromen-2-one (3, 6a-e, 10a-c and 12a-c) derivatives were assessed against VEGFR-2 and demonstrated comparable activities to that of Sorafenib (approved medicine) with compounds 6d and 6b showing the highest binding scores (-9.900 and -9.819 kcal/mol, respectively). The cytotoxicity of the most active thiazole derivatives 6d, 6b, 6c, 10c and 10a were investigated for their human breast cancer (MCF-7) cell line and normal cell line LLC-Mk2 using MTT assay and Sorafenib as the reference drug. The results revealed that compounds 6d and 6b exhibited greater anticancer activities (IC50 = 10.5 ± 0.71 and 11.2 ± 0.80 µM, respectively) than the Sorafenib reference drug (IC50 = 5.10 ± 0.49 µM). Therefore, the present study demonstrated that thiazolyl coumarins are potential (VEGFR-2) inhibitors and pave the way for the synthesis of additional libraries based on the reported scaffold, which could eventually lead to the development of efficient treatment for breast cancer.


Assuntos
Antineoplásicos , Neoplasias da Mama , Cumarínicos , Receptor 2 de Fatores de Crescimento do Endotélio Vascular , Feminino , Humanos , Antineoplásicos/química , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células , Cumarínicos/química , Cumarínicos/farmacologia , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Simulação de Acoplamento Molecular , Estrutura Molecular , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Sorafenibe/farmacologia , Relação Estrutura-Atividade , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
16.
Molecules ; 28(2)2023 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-36677692

RESUMO

Toll-like receptor 7 (TLR7) is a class of pattern recognition receptors (PRRs) recognizing the pathogen-associated elements and damage and as such is a major player in the innate immune system. TLR7 triggers the release of pro-inflammatory cytokines or type-I interferons (IFN), which is essential for immunoregulation. Increasing reports also highlight that the abnormal activation of endosomal TLR7 is implicated in various immune-related diseases, carcinogenesis as well as the proliferation of human immunodeficiency virus (HIV). Hence, the design and development of potent and selective TLR7 antagonists based on small molecules or oligonucleotides may offer new tools for the prevention and management of such diseases. In this review, we offer an updated overview of the main structural features and therapeutic potential of small-molecule antagonists of TLR7. Various heterocyclic scaffolds targeting TLR7 binding sites are presented: pyrazoloquinoxaline, quinazoline, purine, imidazopyridine, pyridone, benzanilide, pyrazolopyrimidine/pyridine, benzoxazole, indazole, indole, and quinoline. Additionally, their structure-activity relationships (SAR) studies associated with biological activities and protein binding modes are introduced.


Assuntos
Interferon Tipo I , Receptor 7 Toll-Like , Humanos , Citocinas/metabolismo , Relação Estrutura-Atividade , Receptor 7 Toll-Like/antagonistas & inibidores
17.
Molecules ; 28(2)2023 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-36677825

RESUMO

SARS-CoV-2 nsp14 guanine-N7-methyltransferase plays an important role in the viral RNA translation process by catalyzing the transfer of a methyl group from S-adenosyl-methionine (SAM) to viral mRNA cap. We report a structure-guided design and synthesis of 3-(adenosylthio)benzoic acid derivatives as nsp14 methyltransferase inhibitors resulting in compound 5p with subnanomolar inhibitory activity and improved cell membrane permeability in comparison with the parent inhibitor. Compound 5p acts as a bisubstrate inhibitor targeting both SAM and mRNA-binding pockets of nsp14. While the selectivity of 3-(adenosylthio)benzoic acid derivatives against human glycine N-methyltransferase was not improved, the discovery of phenyl-substituted analogs 5p,t may contribute to further development of SARS-CoV-2 nsp14 bisubstrate inhibitors.


Assuntos
Antivirais , Metiltransferases , SARS-CoV-2 , Metilação , Metiltransferases/antagonistas & inibidores , RNA Mensageiro/genética , RNA Viral/genética , S-Adenosilmetionina/química , SARS-CoV-2/efeitos dos fármacos , Proteínas não Estruturais Virais/metabolismo , Antivirais/farmacologia
18.
J Transl Med ; 21(1): 31, 2023 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-36650543

RESUMO

NOC2 like nucleolar associated transcriptional repressor (NOC2L) was recently identified as a novel inhibitor of histone acetyltransferase (INHAT). NOC2L is found to have two INHAT function domains and regulates histone acetylation in a histone deacetylases (HDAC) independent manner, which is distinct from other INHATs. In this review, we summarize the biological function of NOC2L in histone acetylation regulation, P53-mediated transcription, ribosome RNA processing, certain development events and carcinogenesis. We propose that NOC2L may be explored as a potential biomarker and a therapeutic target in clinical practice.


Assuntos
Histona Acetiltransferases , Histonas , Proteínas Repressoras , Acetilação , Histona Acetiltransferases/antagonistas & inibidores , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Proteínas Repressoras/genética , Fatores de Transcrição/genética
19.
Skin Res Technol ; 29(1): e13275, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36704884

RESUMO

BACKGROUND: Sensitive skin is a subjective cutaneous hyper-reactivity that occurs in response to various innocuous stimuli. Keratinocytes have recently been shown to participate in sensory transduction by releasing many neuroactive molecules that bind to intra-epidermal free nerve endings and modulate nociception. In the literature, the characterization of these interactions has been based on the co-culture of keratinocyte and mammalian-origin neuronal cell lines. In this study, we established an in vitro model based on a co-culture of primary human keratinocytes and differentiated SH-SY5Y cells, a human neuronal cell line. METHODS: Human epidermal keratinocytes and SH-SY5Y cells were monocultured and co-cultured. Changes in calcium influx, substance P, inflammatory cytokines, and neuropeptides between the monoculture and co-culture groups treated with capsaicin only and capsaicin with transient receptor potential channel vanilloid subfamily member 1 (TRPV1) antagonist, trans-4-tert-butylcyclohexanol (TTBC), together. In addition, the difference in stinging sensation was evaluated by applying it to the volunteers. RESULTS: When SH-SY5Y cells were co-cultured with keratinocytes, they had no significant effect on axonal development. Substance P was also released after capsaicin treatment and reduced by TTBC under co-culture conditions. Moreover, the expression of inflammatory cytokines and neuropeptides was significantly increased in co-cultured keratinocytes compared to that under monoculture conditions. In addition, the stinging sensation was significantly induced after the application of capsaicin in vivo and was relieved after the application of the TRPV1 antagonist. CONCLUSION: We demonstrated that the novel co-culture model is functionally valid through capsaicin and TRPV1 antagonist. We also confirmed that TTBC could be used for the treatment of sensitive skin through a co-culture model and in vivo tests. This co-culture model of keratinocytes and SH-SY5Y cells may be useful in vitro alternatives for studying the close communication between keratinocytes and neuronal cells and for screening therapeutic drugs for sensitive skin.


Assuntos
Neuroblastoma , Neuropeptídeos , Canais de Cátion TRPV , Animais , Humanos , Capsaicina/farmacologia , Linhagem Celular , Técnicas de Cocultura , Citocinas/metabolismo , Queratinócitos/metabolismo , Neuroblastoma/metabolismo , Neuropeptídeos/metabolismo , Substância P/metabolismo , Canais de Cátion TRPV/antagonistas & inibidores
20.
Phytomedicine ; 110: 154637, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36610353

RESUMO

BACKGROUND: Non-peptide thrombopoietin receptor (TPOR) agonists are promising therapies for the mitigation and treatment of thrombocytopenia. However, only few agents are available as safe and effective for stimulating platelet production for thrombocytopenic patients in the clinic. PURPOSE: This study aimed to develop a novel small molecule TPOR agonist and investigate its underlying regulation of function in megakaryocytes (MKs) differentiation and thrombopoiesis. METHODS: A potential active compound that promotes MKs differentiation and thrombopoiesis was obtained by machine learning (ML). Meanwhile, the effect was verified in zebrafish model, HEL and Meg-01 cells. Next, the key regulatory target was identified by Drug Affinity Responsive Target Stabilization Assay (DARTS), Cellular Thermal Shift Assay (CETSA), and molecular simulation experiments. After that, RNA-sequencing (RNA-seq) was used to further confirm the associated pathways and evaluate the gene expression induced during MK differentiation. In vivo, irradiation (IR) mice, C57BL/6N-TPORem1cyagen (Tpor-/-) mice were constructed by CRISPR/Cas9 technology to examine the therapeutic effect of TMEA on thrombocytopenia. RESULTS: A natural chemical-structure small molecule TMEA was predicted to be a potential active compound based on ML. Obvious phenotypes of MKs differentiation were observed by TMEA induction in zebrafish model and TMEA could increase co-expression of CD41/CD42b, DNA content, and promote polyploidization and maturation of MKs in HEL and Meg-01 cells. Mechanically, TMEA could bind with TPOR protein and further regulate the PI3K/AKT/mTOR/P70S6K and MEK/ERK signal pathways. In vivo, TMEA evidently promoted platelet regeneration in mice with radiation-induced thrombocytopenia but had no effect on Tpor-/- and C57BL/6 (WT) mice. CONCLUSION: TMEA could serve as a novel TPOR agonist to promote MKs differentiation and thrombopoiesis via mTOR and ERK signaling and could potentially be created as a promising new drug to treat thrombocytopenia.


Assuntos
Trombocitopenia , Trombopoese , Animais , Camundongos , Diferenciação Celular , Megacariócitos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Trombocitopenia/tratamento farmacológico , Trombocitopenia/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Peixe-Zebra/metabolismo , Sistema de Sinalização das MAP Quinases , Receptores de Trombopoetina/antagonistas & inibidores
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