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1.
Phys Chem Chem Phys ; 23(36): 20230-20246, 2021 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-34474468

RESUMO

Research on action selectivity between CYP1A1 and CYP1B1 is particularly valuable for cancer chemoprevention and chemotherapy. However, they share a very close similarity in their ligand-binding pockets that α-naphthoflavone (ANF) is the co-crystal ligand for both isoforms, which poses a major challenge in revealing their selectivity mechanism. Therefore, three selective CYP1B1 inhibitors derived from ANF were selected to illustrate the structural basis for the selectivity between the two isoforms via a comprehensive computational strategy. It was found that the sustainability of the π-π stacking interactions with the phenylalanine residues of the two isoforms, namely, Phe123, Phe224, and Phe258 for CYP1A1, and Phe134, Phe231, and Phe268 for CYP1B1, played a crucial role in determining the selectivity of ligands with a classic aromatic conjugation system like ANF and its derivatives for CYP1B1 versus CYP1A1. Of note, the structural flexibility of the corresponding protein domains mainly orchestrated the sustainability of the corresponding π-π stacking interactions, thereby determining the binding selectivity. Therefore, the structure modification of naphthoflavone lead compounds into preferable binding configurations to satisfy the π-π stacking interactions of the key phenylalanine residues within CYP1B1 would be an inspiring strategy devised to improve the inhibitory selectivity towards CYP1B1. Collectively, this study revealed valuable insight into understanding the selective mechanism between CYP1A1 and CYP1B1 from the perspective of structural flexibility, which sheds light on the future rational design of CYP1B1 selective inhibitors.


Assuntos
Benzoflavonas/farmacologia , Citocromo P-450 CYP1A1/antagonistas & inibidores , Citocromo P-450 CYP1B1/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Benzoflavonas/química , Citocromo P-450 CYP1A1/química , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1B1/química , Citocromo P-450 CYP1B1/metabolismo , Inibidores Enzimáticos/química , Humanos , Simulação de Dinâmica Molecular , Estrutura Molecular
2.
Bioorg Chem ; 116: 105295, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34455300

RESUMO

We previously discovered extrahepatic cytochrome P450 1B1 (CYP1B1) degraders able to overcome drug resistance toward docetaxel using a PROTACs technology, however, the underexplored structure activity relationships and poor water solubility posed a major hurdle in the development of CYP1B1 degraders. Herein, continuous efforts are made to develop more promising α-naphthoflavone (ANF)-derived chimeras for degrading CYP1B1. Guided by the strongest ANF-derived CYP1B1 degrader 3a we ever reported, 17 ANF analogues are designed and synthesized to evaluate the CYP1B1 degradation and resultant resistance reversal. In degrading CYP1B1 and sensitizing drug resistance, 4d with a 1, 5-cis triazole coupling mode at (C3') of B ring of ANF exhibited the similar potency as 3a carrying a 1, 4-trans triazole fragment at (C4') of B ring, but more obvious selectivity of 4d toward CYP1B1 over CYP1A2 is observed. When an oxygen was inserted into the linker of 4d, 4f demonstrated better water solubility, a more potent ability in degrading CYP1B1 and reversing drug resistance, and a promising selectivity. Collectively, a substitution position, an alkyne-azide cyclization and a liker type significantly affect the ability of ANF-thalidomide conjugates in eliminating drug resistance of CYP1B1-expressing DU145 (DU145/CY) cells to docetaxel via targeted CYP1B1 degradation.


Assuntos
Antineoplásicos/farmacologia , Benzoflavonas/farmacologia , Citocromo P-450 CYP1B1/antagonistas & inibidores , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias da Próstata/tratamento farmacológico , Antineoplásicos/síntese química , Antineoplásicos/química , Benzoflavonas/síntese química , Benzoflavonas/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citocromo P-450 CYP1B1/metabolismo , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Masculino , Estrutura Molecular , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Solubilidade , Relação Estrutura-Atividade
3.
Biol Pharm Bull ; 44(4): 579-584, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33790108

RESUMO

The inhibitory and stimulatory effects of steroid hormones and related compounds on the hydroxylation activity at the 6ß-position of two steroid hormones, progesterone and testosterone, by CYP3A4, polymorphically expressed CYP3A5, and fetal CYP3A7 were compared to clarify the catalytic properties of the predominant forms of the human CYP3A subfamily. Hydroxylation activities of progesterone and testosterone by CYP3A4, CYP3A5, and CYP3A7 were estimated using HPLC. The Michaelis constants (Km) for progesterone 6ß-hydroxylation by CYP3A5 were markedly decreased in the presence of dehydroepiandrosterone (DHEA) and α-naphthoflavone (ANF), whereas progesterone and DHEA competitively inhibited testosterone 6ß-hydroxylation mediated by CYP3A4, and progesterone competitively inhibited CYP3A5-mediated activity, which was weaker than that for CYP3A4. ANF noncompetitively inhibited testosterone 6ß-hydroxylation mediated by both CYP3A4 and CYP3A5. Progesterone and testosterone 6ß-hydroxylation mediated by CYP3A7 was inhibited or unaffected by DHEA, pregnenolone, and ANF. These results suggested that DHEA and ANF stimulated progesterone 6ß-hydroxylation by CYP3A5 but not by CYP3A4 and CYP3A7; however, progesterone, DHEA, and ANF inhibited testosterone 6ß-hydroxylation mediated by all CYP3A subfamily members. The inhibitory/stimulatory pattern of steroid-steroid interactions is different among CYP3A subfamily members and CYP3A5 is the most sensitive in terms of activation among the CYP3A subfamily members investigated.


Assuntos
Benzoflavonas/farmacologia , Citocromo P-450 CYP3A/metabolismo , Esteroides/farmacologia , Catálise , Citocromo P-450 CYP3A/genética , Escherichia coli/genética , Hidroxilação/efeitos dos fármacos
4.
Eur J Med Chem ; 209: 112895, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33069055

RESUMO

Cytochrome P450 1B1 (CYP1B1) has been well validated as an attractive target for cancer prevention and drug resistance reversal. In continuation of our interest in this area, herein, a set of forty-six 6,7,10-trimethoxy-α-naphthoflavone derivatives varying in B ring was synthesized and screened against CYP1 enzymes, leading to the identification of fluorine-containing compound 15i as the most potent and selective CYP1B1 inhibitor (IC50 value of 0.07 nM), being 84-fold more potent than that of the template molecule ANF. Alternatively, the amino-substituted derivative 13h not only possessed a potent inhibitory effect on CYP1B1 (IC50 value of 0.98 nM), but also had a substantially increased water solubility as compared with the lead ANF (311 µg/mL for 13h and <5 µg/mL for ANF). The current study expanded the structural diversity of CYP1B1 inhibitors, and compound 13h could be considered as a promising starting point with great potential for further studies.


Assuntos
Benzoflavonas/química , Benzoflavonas/farmacologia , Citocromo P-450 CYP1B1/antagonistas & inibidores , Inibidores das Enzimas do Citocromo P-450/química , Inibidores das Enzimas do Citocromo P-450/farmacologia , Citocromo P-450 CYP1B1/metabolismo , Descoberta de Drogas , Humanos , Simulação de Acoplamento Molecular , Neoplasias/tratamento farmacológico , Solubilidade , Água/química
5.
Arch Biochem Biophys ; 698: 108677, 2021 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-33197431

RESUMO

We investigate the effect of the alcohol-induced increase in the content of CYP2E1 in human liver microsomes (HLM) on the function of CYP3A4. Membrane incorporation of the purified CYP2E1 into HLM considerably increases the rate of metabolism of 7-benzyloxyquinoline (BQ) and attenuates the homotropic cooperativity observed with this CYP3A4-specific substrate. It also eliminates the activating effect of α-naphthoflavone (ANF) seen in some HLM samples. To probe the physiological relevance of these effects, we compared three pooled preparations of HLM from normal donors (HLM-N) with a pooled preparation from ten heavy alcohol consumers (HLM-A). The composition of the P450 pool in all samples was characterized by the mass-spectrometric determination of 11 cytochrome P450 species. The fractional content of CYP2E1 in HLM-A was from 2.0 to 3.4 times higher than in HLM-N. In contrast, the content of CYP3A4 in HLM-A was the lowest among all samples. Despite that, HLM-A exhibited a much higher metabolism rate and a lower homotropic cooperativity with BQ, similar to CYP2E1-enriched HLM-N. To substantiate the involvement of interactions between CYP2E1 and CYP3A4 in these effects, we probed hetero-association of these proteins in CYP3A4-containing Supersomes™ with a technique employing CYP2E1 labeled with BODIPY-618 maleimide. These experiments evinced the interactions between the two enzymes and revealed an inhibitory effect of ANF on their association. Our results demonstrate that the functional properties of CYP3A4 are fundamentally dependent on the composition of the cytochrome P450 ensemble and suggest a possible impact of chronic alcohol exposure on the pharmacokinetics of drugs metabolized by CYP3A4.


Assuntos
Citocromo P-450 CYP2E1/metabolismo , Citocromo P-450 CYP3A/metabolismo , Etanol/toxicidade , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Sequência de Aminoácidos , Amitriptilina/metabolismo , Benzoflavonas/farmacologia , Citocromo P-450 CYP2E1/análise , Citocromo P-450 CYP3A/análise , Ativadores de Enzimas/farmacologia , Feminino , Humanos , Ivermectina/metabolismo , Masculino , Midazolam/metabolismo , Nitrofenóis/metabolismo , Quinolinas/metabolismo
6.
Eur J Med Chem ; 189: 112028, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31945665

RESUMO

Extrahepatic cytochrome P450 1B1 (CYP1B1), which is highly expressed in various tumors, is an attractive and potential target for cancer prevention, therapy, and reversal of drug resistance. CYP1B1 inhibition is the current predominant therapeutic paradigm to treating CYP1B1-mediated malignancy, but therapeutic effect has little success. Herein, we reported CYP1B1 degradation in place of CYP1B1 inhibition for reversing drug resistance toward docetaxel in CYP1B1-overexpressing prostate cancer cell line DU145 using a PROTAC strategy. Replacing chlorine atom of a CYP1B1 selective inhibitor we found previously with ethynyl, we got the resulting α-naphthoflavone derivative 5 which kept strong inhibition against CYP1B1 (IC50 = 0.4 ± 0.2 nM) and high selectivity. Coupling of 5 with thalidomide derivatives of varying chain lengths afforded conjugates 6A-Dvia click reaction. In vitro cell-based assay indicated that 6C was more effective in eliminating drug resistance of CYP1B1-overexpressed DU145 cells compared with other analogues. Western blotting analysis showed CYP1B1 degradation was one main reason for the reversal of drug resistance to docetaxel and the effect was obtained in a concentration-dependent manner. This work is the first attempt to overcome CYP1B1-mediated drug resistance via CYP1B1 degradation instead of CYP1B1 inhibition, which could provide a new direction toward eliminating drug resistance.


Assuntos
Benzoflavonas/farmacologia , Citocromo P-450 CYP1B1/antagonistas & inibidores , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Talidomida/análogos & derivados , Talidomida/farmacologia , Benzoflavonas/síntese química , Linhagem Celular Tumoral , Citocromo P-450 CYP1B1/metabolismo , Docetaxel/farmacologia , Desenho de Fármacos , Humanos , Proteólise , Talidomida/síntese química
7.
Eur J Med Chem ; 187: 111938, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31830634

RESUMO

Cytochrome P450 1B1(CYP1B1) has been recognized as an important target for cancer prevention and drug resistance reversal. In order to obtain potent and selective CYP1B1 inhibitors, a series of forty-one α-naphthoflavone (ANF) derivatives were synthesized, characterized, and evaluated for CYP1B1, CYP1A1 and CYP1A2 inhibitory activities. A closure look into the structure-activity relationship for the inhibitory effects on CYP1B1 indicated that modification of the C ring of ANF would decrease the CYP1B1 inhibitory potency, while incorporation of substituent(s) into the different positions of the B ring yielded analogues with varying CYP1B1 inhibitory capacity. Among these derivatives, compounds 9e and 9j were identified as the most potent two selective CYP1B1 inhibitors with IC50 values of 0.49 and 0.52 nM, respectively, which were 10-fold more potent than the lead compound ANF. In addition, molecular docking and a reasonable 3D-QSAR (three-dimensional quantitative structure-activity relationship) study were performed to provide a better understanding of the key structural features influencing the CYP1B1 inhibitory activity. The results achieved in this study would lay a foundation for future development of selective, potent, low-toxic and water-soluble CYP1B1 inhibitors.


Assuntos
Benzoflavonas/farmacologia , Citocromo P-450 CYP1B1/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Benzoflavonas/síntese química , Benzoflavonas/química , Citocromo P-450 CYP1B1/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Proteínas Recombinantes/metabolismo , Relação Estrutura-Atividade
8.
Front Biosci (Elite Ed) ; 12(1): 79-94, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31585870

RESUMO

Vascular dementia (VaD) refers to the loss of reasoning, planning, judgment, and memory as a result of reduced blood flow, inflammation, activation of immune cells, and neurodegenerative processes. Here, we tested the preventive effects of alpha-naphthoflavone (ANF) on a model of vascular dementia induced by four-vessel occlusion (4VO) in rats. Animals were treated orally for 5 consecutive days with 10 mg/kg of donepezil (a traditional therapeutic drug) and 40 and 80 mg/kg ANF. The treatment with ANF exerted effects that were similar to those induced by donepezil. This included prevention of cognitive impairment, alterations in the plasma levels of homocysteine and nitrate and the activity of acetylcholinesterase and myeloperoxidase and the tissue level of glutathione, and thiobarbituric acid reactive substances in the brain tissues. These findings suggest that herbal-derived ANF is as effective as traditional drugs in vascular dementia.


Assuntos
Benzoflavonas/uso terapêutico , Demência Vascular/prevenção & controle , Animais , Benzoflavonas/farmacologia , Demência Vascular/sangue , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Teste de Labirinto em Cruz Elevado , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Teste do Labirinto Aquático de Morris/efeitos dos fármacos , Ratos Wistar , Comportamento Social
9.
Biomed Pharmacother ; 118: 109287, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31401392

RESUMO

Non-alcoholic fatty liver disease (NAFLD) is a chronic liver disease. The literature suggests that the aryl hydrocarbon receptor (AHR) may be a key player in the pathogenesis of NAFLD, and it can modulate the synthesis of cytochrome P450 1A1 (CYP1A1) and tumor necrosis factor-α (TNF-α). Previous studies have shown that CYP1A1 is a key enzyme of oxidative stress, TNF-α is involved in the formation of insulin resistance (IR), oxidative stress and insulin resistance are the key factors for the formation of NAFLD. Therefore, it can be said that AHR may participate in contributing to NAFLD by regulating CYP1A1 and TNF-α. Alpha-naphthoflavone (ANF) is an effective AHR inhibitor. The present study was designed to explore the hepatoprotective effect of ANF in high fat diet (HFD)-induced NAFLD mice and oleic acid (OA)-treated HepG2 hepatocytes. Mice were fed HFD to induce NAFLD, HepG2 cells were exposed to OA to induce hepatocyte injury, and ANF significantly reduced mouse and cellular liver damage compared to the HFD-induced NAFLD and OA-treated HepG2 hepatocytes. ANF treatment reduces liver damage by reducing ROS and IR, the data show that ANF inhibits the expression of AHR, CYP1A1 and TNF-α in NAFLD. Taken together, these findings show that ANF alleviate NAFLD via regulation of AHR/CYP1A1 and AHR/TNF-α pathways, which may have potential for further development as novel therapeutic agents for NAFLD.


Assuntos
Benzoflavonas/uso terapêutico , Hepatócitos/patologia , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Animais , Benzoflavonas/farmacologia , Catalase/metabolismo , Proliferação de Células/efeitos dos fármacos , Dieta Hiperlipídica , Glicogênio/metabolismo , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Humanos , Resistência à Insulina , Gotículas Lipídicas/efeitos dos fármacos , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Malondialdeído/metabolismo , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/patologia , Ácido Oleico , Estresse Oxidativo/efeitos dos fármacos , Superóxido Dismutase/metabolismo
10.
Nat Commun ; 10(1): 1125, 2019 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-30850589

RESUMO

Whether tobacco carcinogens enable exposed cells immune escape resulting in carcinogenesis, and why patients who smoke respond better to immunotherapies than non-smokers, remains poorly understood. Here we report that cigarette smoke and the carcinogen benzo(a)pyrene (BaP) induce PD-L1 expression on lung epithelial cells in vitro and in vivo, which is mediated by aryl hydrocarbon receptor (AhR). Anti-PD-L1 antibody or deficiency in AhR significantly suppresses BaP-induced lung cancer. In 37 patients treated with anti-PD-1 antibody pembrolizumab, 13/16 (81.3%) patients who achieve partial response or stable disease express high levels of AhR, whereas 12/16 (75%) patients with progression disease exhibit low levels of AhR in tumor tissues. AhR inhibitors exert significant antitumor activity and synergize with anti-PD-L1 antibody in lung cancer mouse models. These results demonstrate that tobacco smoke enables lung epithelial cells to escape from adaptive immunity to promote tumorigenesis, and AhR predicts the response to immunotherapy and represents an attractive therapeutic target.


Assuntos
Anticorpos Monoclonais/farmacologia , Antígeno B7-H1/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Receptores de Hidrocarboneto Arílico/genética , Carcinoma de Pequenas Células do Pulmão/genética , Tabaco/toxicidade , Animais , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/imunologia , Benzo(a)pireno/toxicidade , Benzoflavonas/farmacologia , Carcinógenos/toxicidade , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Humanos , Imunoterapia/métodos , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Neoplasias Pulmonares/etiologia , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/terapia , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Hidrocarboneto Arílico/antagonistas & inibidores , Receptores de Hidrocarboneto Arílico/imunologia , Transdução de Sinais , Carcinoma de Pequenas Células do Pulmão/etiologia , Carcinoma de Pequenas Células do Pulmão/mortalidade , Carcinoma de Pequenas Células do Pulmão/terapia , Fumar/efeitos adversos , Análise de Sobrevida , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Xenobiotica ; 49(9): 1015-1024, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30272491

RESUMO

Cytochrome P450 (CYP) enzymes constitute an essential xenobiotic metabolizing system that regulates the elimination of lipophilic compounds from the body. Convenient and affordable assays for CYP enzymes are important for assessing these metabolic pathways. In this study, 10 novel profluorescent coumarin derivatives with various substitutions at carbons 3, 6 and 7 were developed. Molecular modeling indicated that 3-phenylcoumarin offers an excellent scaffold for the development of selective substrate compounds for various human CYP forms, as they could be metabolized to fluorescent 7-hydroxycoumarin derivatives. Oxidation of profluorescent coumarin derivatives to fluorescent metabolites by 13 important human liver xenobiotic-metabolizing CYP forms was determined by enzyme kinetic assays. Four of the coumarin derivatives were converted to fluorescent metabolites by CYP1 family enzymes, with 6-methoxy-3-(4-trifluoromethylphenyl)coumarin being oxidized selectively by CYP1A2 in human liver microsomes. Another set of four compounds were metabolized by CYP2A6 and CYP1 enzymes. 7-Methoxy-3-(3-methoxyphenyl)coumarin was oxidized efficiently by CYP2C19 and CYP2D6 in a non-selective fashion. The advantages of the novel substrates were (1) an excellent signal-to-background ratio, (2) selectivity for CYP1 forms, and (3) convenient multiwell plate measurement, allowing for precise determination of potential inhibitors of important human hepatic forms CYP1A2, CYP2C19 and CYP2D6.


Assuntos
Cumarínicos/química , Cumarínicos/metabolismo , Sistema Enzimático do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/metabolismo , Benzoflavonas/metabolismo , Benzoflavonas/farmacologia , Cumarínicos/síntese química , Inibidores das Enzimas do Citocromo P-450/metabolismo , Inibidores das Enzimas do Citocromo P-450/farmacologia , Sistema Enzimático do Citocromo P-450/genética , Fluorescência , Humanos , Inativação Metabólica , Cinética , Microssomos Hepáticos/metabolismo , Modelos Moleculares , Simulação de Acoplamento Molecular , Oxirredução
12.
Bioorg Chem ; 78: 178-184, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29574302

RESUMO

In this work, 17α-methyltestosterone was effectively hydroxylated by Absidia coerulea KCh 93, Syncephalastrum racemosum KCh 105 and Chaetomium sp. KCh 6651. A. coerulea KCh 93 afforded 6ß-, 12ß-, 7α-, 11α-, 15α-hydroxy derivatives with 44%, 29%, 6%, 5% and 9% yields, respectively. S. racemosum KCh 105 afforded 7α-, 15α- and 11α-hydroxy derivatives with yields of 45%, 19% and 17%, respectively. Chaetomium sp. KCh 6651 afforded 15α-, 11α-, 7α-, 6ß-, 9α-, 14α-hydroxy and 6ß,14α-dihydroxy derivatives with yields of 31%, 20%, 16%, 7%, 5%, 7% and 4%, respectively. 14α-Hydroxy and 6ß,14α-dihydroxy derivatives were determined as new compounds. Effect of various sources of nitrogen and carbon in the media on biotransformations were tested, however did not affect the degree of substrate conversion or the composition of the products formed. The addition of α- or ß-naphthoflavones inhibited 17α-methyltestosterone hydroxylation but did not change the percentage composition of the resulting products.


Assuntos
Benzoflavonas/farmacologia , Inibidores Enzimáticos/farmacologia , Metiltestosterona/antagonistas & inibidores , Oxigenases de Função Mista/antagonistas & inibidores , beta-Naftoflavona/farmacologia , Absidia/enzimologia , Benzoflavonas/síntese química , Benzoflavonas/química , Chaetomium/enzimologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Metiltestosterona/química , Metiltestosterona/metabolismo , Oxigenases de Função Mista/metabolismo , Estrutura Molecular , Mucorales/enzimologia , Relação Estrutura-Atividade , beta-Naftoflavona/síntese química , beta-Naftoflavona/química
13.
Med Chem ; 14(4): 322-332, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29332599

RESUMO

BACKGROUND: Although the expression levels of many P450s differ between tumour and corresponding normal tissue, CYP1B1 is one of the few CYP subfamilies which is significantly and consistently overexpressed in tumours. CYP1B1 has been shown to be active within tumours and is capable of metabolising a structurally diverse range of anticancer drugs. Because of this, and its role in the activation of procarcinogens, CYP1B1 is seen as an important target for anticancer drug development. OBJECTIVE: To synthesise a series of chalcone derivatives based on the chemopreventative agent DMU-135 and investigate their antiproliferative activities in human breast cancer cell lines which express CYP1B1 and CYP1A1. METHOD: A series of chalcones were synthesised in yields of 43-94% using the Claisen-Schmidt condensation reaction. These were screened using a MTT assay against a panel of breast cancer cell lines which have been characterised for CYP1 expression. RESULT: A number of derivatives showed promising antiproliferative activities in human breast cancer cell lines which express CYP1B1 and CYP1A1, while showing significantly lower toxicity towards a non-tumour breast cell line with no CYP expression. Experiments using the CYP1 inhibitors acacetin and α-naphthoflavone provided supporting evidence for the involvement of CYP1 enzymes in the bioactivation of these compounds. CONCLUSION: Chalcones show promise as anticancer agents with evidence suggesting that CYP1 activation of these compounds may be involved.


Assuntos
Antineoplásicos/farmacologia , Chalcona/análogos & derivados , Inibidores das Enzimas do Citocromo P-450/farmacologia , Pró-Fármacos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/toxicidade , Benzoflavonas/farmacologia , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Chalcona/síntese química , Chalcona/química , Chalcona/farmacologia , Chalcona/toxicidade , Citocromo P-450 CYP1A1/antagonistas & inibidores , Citocromo P-450 CYP1B1/antagonistas & inibidores , Inibidores das Enzimas do Citocromo P-450/síntese química , Inibidores das Enzimas do Citocromo P-450/química , Inibidores das Enzimas do Citocromo P-450/toxicidade , Flavonas/farmacologia , Humanos , Estrutura Molecular , Pró-Fármacos/síntese química , Pró-Fármacos/química , Pró-Fármacos/toxicidade
14.
Toxicol Appl Pharmacol ; 338: 73-82, 2018 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-29146461

RESUMO

Infertility is a global health problem with an estimated incidence of 15%. Exposure to chemicals is a potential causal factor, and there is a lack of studies examining the effects on female germ cells. Here, we have studied the impact of different aryl hydrocarbon receptor (AHR) modulators on human ovarian follicles using a human ovarian tissue culture model. Expression of AHR was analyzed in tissue samples, and effects of the selected ligands resveratrol (RSVL), 6-formylindolo(3,2-b)carbazole (FICZ), and alpha-naphthoflavone (aNF) on AHR transactivation studied in a granulosa cell tumor line. Cortical human ovarian tissue containing preantral follicles was exposed to the ligands or vehicle (dimethylsulfoxide, DMSO) for seven days in vitro. Follicle growth was assessed by counting and measuring follicles from serial tissue sections, cell death quantified using in situ Terminal deoxynucleotidyl transferase dUTP Nick-End Labeling (TUNEL) assay, and steroid hormone production measured using a newly developed ultra-performance liquid chromatography method. AHR was expressed in all donated ovarian tissue samples. FICZ induced AHR transactivation in the granulosa cell line while aNF antagonised it. Compared to DMSO control, FICZ had no effect on follicles in culture, RSVL increased the proportion of growing follicles, and aNF increased cell death, disrupted growth of secondary follicles, increased testosterone, and reduced estradiol levels. We conclude that RSVL supports and aNF disrupts growth of human ovarian follicles in culture. We further conclude that the human ovarian tissue culture model is suitable for studying effects of chemicals on follicular biology.


Assuntos
Benzoflavonas/farmacologia , Folículo Ovariano/efeitos dos fármacos , Estilbenos/farmacologia , Adulto , Carbazóis/farmacologia , Morte Celular/efeitos dos fármacos , Feminino , Humanos , Marcação In Situ das Extremidades Cortadas , Folículo Ovariano/crescimento & desenvolvimento , Receptores de Hidrocarboneto Arílico/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/genética , Resveratrol , Técnicas de Cultura de Tecidos
15.
Arch Toxicol ; 92(2): 907-919, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29094188

RESUMO

Daily exposure to low doses of 3-methylcholanthrene (3MC) during the pubertal period in rats disrupts both follicular growth and ovulation. Thus, to provide new insights into the toxicity mechanism of 3MC in the ovary, here we investigated the effect of daily exposure to 3MC on selected ovarian genes, the role of the aryl hydrocarbon receptor (AhR) and the level of epigenetic remodeling of histone post-transcriptional modifications. Immature rats were daily injected with 3MC (0.1 or 1 mg/kg) and mRNA expression of genes involved in different ovarian processes were evaluated. Of the 29 genes studied, 18 were up-regulated, five were down-regulated and six were not altered. To assess whether AhR was involved in these changes, we used the chromatin immunoprecipitation assay. 3MC increased AhR binding to promoter regions of genes involved in Notch signaling (Hes1, Jag1), activation of primordial follicles (Cdk2), cell adhesion (Icam1), stress and tumor progression (Dnajb6), apoptosis (Bax, Caspase-9) and expression of growth and transcription factors (Igf2, Sp1). Studying the trimethylation and acetylation of histone 3 (H3K4me3 and H3K9Ac, respectively) of these genes, we found that 3MC increased H3K4me3 in Cyp1a1, Jag1, Dnajb6, Igf2, Notch2, Adamts1, Bax and Caspase-9, and H3K9Ac in Cyp1a1, Jag1, Cdk2, Dnajb6, Igf2, Icam1, and Sp1. Co-treatment with α-naphthoflavone (αNF), a specific antagonist of AhR, prevented almost every 3MC-induced changes. Despite the low dose used in these experiments, daily exposure to 3MC induced changes in both gene expression and epigenomic remodeling, which may lead to premature ovarian failure.


Assuntos
Benzoflavonas/farmacologia , Metilcolantreno/toxicidade , Folículo Ovariano/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/metabolismo , Acetilação , Animais , Imunoprecipitação da Cromatina , Regulação para Baixo , Epigênese Genética , Feminino , Histonas/química , Metilação , Regiões Promotoras Genéticas , Ligação Proteica , Processamento de Proteína Pós-Traducional , Ratos , Ratos Sprague-Dawley , Regulação para Cima
16.
Nutr Res ; 44: 38-50, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28821316

RESUMO

Inhibition of the aryl hydrocarbon receptor (AHR) prevents Western diet-induced obesity and fatty liver in C57Bl/6J (B6) male mice. The AHR is a ligand-activated nuclear receptor that regulates genes involved in xenobiotic metabolism and T-cell differentiation. Here, we tested the hypothesis that AHR antagonism would also prevent obesity and fatty liver in female mice and that B6 mice (higher-affinity AHR) and congenic B6.D2 mice (lower-affinity AHR) would differentially respond to AHR inhibition. Female and male adult B6 and B6.D2 mice were fed control and Western diets with and without α-naphthoflavone (NF), an AHR inhibitor. A nonlinear mixed-model analysis was developed to project asymptote body mass. We found that obesity, adiposity, and liver steatosis were reduced to near control levels in all female and male B6 and B6.D2 experimental groups fed Western diet with NF. However, differences were noted in that female B6.D2 vs B6 mice on Western diet became more obese; and in general, female mice compared with male mice had a greater fat mass to body mass ratio, were less responsive to NF, and had reduced liver steatosis and hepatomegaly. We report that male mice fed Western diet containing NF or CH-223191, another AHR inhibitor, caused reduced mRNA levels of several liver genes involved in metabolism, including Cyp1b1 and Scd1, offering evidence for a possible mechanism by which the AHR regulates obesity. In conclusion, although there are some sex- and Ahr allelic-dependent differences, AHR inhibition prevents obesity and liver steatosis in both males and females regardless of the ligand-binding capacity of the AHR. We also present evidence consistent with the notion that an AHR-CYP1B1-SCD1 axis is involved in obesity, providing potentially convenient and effective targets for treatment.


Assuntos
Benzoflavonas/farmacologia , Fígado Gorduroso/prevenção & controle , Obesidade/prevenção & controle , Receptores de Hidrocarboneto Arílico/antagonistas & inibidores , Adiposidade/efeitos dos fármacos , Animais , Compostos Azo/farmacologia , Citocromo P-450 CYP1B1/genética , Citocromo P-450 CYP1B1/metabolismo , Dieta Ocidental , Feminino , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pirazóis/farmacologia , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismo
17.
Cleft Palate Craniofac J ; 54(2): 216-222, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-27018527

RESUMO

OBJECTIVE: Tocompare the effect of folic acid (FA) and α-naphthoflavone on 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced cleft palate in fetal mice. DESIGN: Pregnant mice were randomly divided into seven groups. The mice treated with corn oil were used as a negative control. The mice in the other six groups were given a single dose of 28 µg/kg TCDD on GD 10 by gavage. For FA treatment, TCDD-treated mice were also dosed with 5, 10, and 15 mg/kg FA on GD 10, while for α-naphthoflavone treatment, the mice received a single dose of 50 µg/kg or 5 mg/kg α-naphthoflavone on GD 10. MAIN OUTCOME MEASURES: Fetal mice palates were imaged using light and scanning electron microscopy on GD 13.5, GD 14.5, and GD 15.5, and cleft palate were recorded on GD 17.5. The expression of guanosine diphosphate dissociation inhibitor (GDI) in fetal mice palate on GD 15.5 was examined by immunohistochemistry. RESULTS: TCDD successfully induced cleft palate. Ten mg/ml FA and 5 mg/ml α-naphthoflavone significantly reduced TCDD-induced cleft palate. FA and α-naphthoflavone partly reduced TCDD-induced cleft palate but did not affect the expression of Rho GDI. CONCLUSIONS: FA and α-naphthoflavone may reduce the generation of reactive oxygen species, inhibit MEE apoptosis through anti-oxidation, and increase filopodia and MEE movement. This may result in restoration of the ultrastructure of the palatal surface to a normal state, leading to the fusion and formation of complete palate in TCDD-treated fetal mice.


Assuntos
Anormalidades Induzidas por Medicamentos/prevenção & controle , Benzoflavonas/farmacologia , Fissura Palatina/induzido quimicamente , Fissura Palatina/prevenção & controle , Ácido Fólico/farmacologia , Dibenzodioxinas Policloradas/toxicidade , Animais , Feminino , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Varredura , Gravidez , Distribuição Aleatória
18.
Cell Reprogram ; 18(3): 154-61, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27253627

RESUMO

Primordial germ cells (PGCs) are the undifferentiated progenitors of gametes. Germline competent PGCs can be developed as a cell-based system for genetic modification in chickens, which provides a valuable tool for transgenic technology with both research and industrial applications. This implies manipulation of PGCs, which, in recent years, encouraged a lot of research focused on the study of PGCs and the way of improving their culture. The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that besides mediating toxic responses to environmental contaminants plays pivotal physiological roles in various biological processes. Since a novel compound that acts as an antagonist of this receptor has been reported to promote expansion of hematopoietic stem cells, we conducted the present study with the aim of determining whether addition of an established AHR antagonist to the standard culture medium used nowadays for in vitro chicken PGCs culture improves ex vivo expansion. We have found that addition of α-naphthoflavone in culture medium promotes the amplification of undifferentiated cells and that this effect is exerted by the blockade of AHR action. Our results constitute the first report of the successful use of a readily available AHR antagonist to improve avian PGCs expansion, and they further extend the knowledge of the effects of AHR modulation in undifferentiated cells.


Assuntos
Benzoflavonas/farmacologia , Células Germinativas/citologia , Células Germinativas/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/antagonistas & inibidores , Transdução de Sinais , Animais , Células Cultivadas , Galinhas , Feminino
19.
Toxicology ; 353-354: 58-69, 2016 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-27163632

RESUMO

In the present study, we investigated the effect of 3-methylcholanthrene (3MC) on sexual maturity and the ability of α-naphthoflavone (αNF) to prevent this action. To this end, immature rats were daily injected intraperitoneally with 3MC (0.1 or 1mg/kg) and/or αNF (80mg/kg). Body weight, vaginal opening and estrous cycle were recorded and ovaries were obtained on the day of estrus. Ovarian weight, ovulation rate (measured by the number of oocytes within oviducts), and follicular development (determined by histology) were studied. No differences were found in body weight, ovarian weight, day of vaginal opening, or the establishment of the estrous cycle among the different groups of rats. However, animals treated with 3MC, at both doses, exhibited a lower number of primordial, primary, preantral and antral follicles than controls. Also, 3MC inhibited the ovulation rate and induced an overexpression of both the Cyp1a1 and Cyp1b1 genes, measured by chromatin immunoprecipitation assay. The daily treatment with αNF alone increased the number of follicles in most of the stages analyzed when compared with controls. Moreover, the αNF treatment prevented completely not only the 3MC-induced decrease in all types of follicles but also the 3MC-induced overexpression of Cyp enzymes and the genetic damage in bone marrow cells and oocytes. These results suggest that (i) daily exposure to 3MC during the pubertal period destroys the follicle reserve and alters the ovulation rate; (ii) the 3MC action seems to be mediated by an aryl hydrocarbon receptor-dependent mechanism; (iii) daily administration of αNF has a clear stimulatory action on the ovarian function; and (iv) αNF may prevent both the systemic and gonadal 3MC-induced toxicity.


Assuntos
Benzoflavonas/administração & dosagem , Benzoflavonas/farmacologia , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1B1/genética , Metilcolantreno/toxicidade , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/patologia , Imunoprecipitação da Cromatina , Relação Dose-Resposta a Droga , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Oócitos/efeitos dos fármacos , Oócitos/patologia , Folículo Ovariano/efeitos dos fármacos , Ovulação/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores de Hidrocarboneto Arílico/metabolismo
20.
Biochem Pharmacol ; 110-111: 117-29, 2016 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-27112072

RESUMO

6-Formylindolo[3,2-b]carbazole (FICZ) is a potent aryl hydrocarbon receptor (AHR) agonist that is efficiently metabolized by AHR-regulated cytochrome P4501 enzymes. FICZ is a proposed physiological AHR ligand that induces its own degradation as part of a regulatory negative feedback loop. In vitro studies in cells show that CYP1 inhibition in the presence of FICZ results in enhanced AHR activation, suggesting that FICZ accumulates in the cell when its metabolism is blocked. We used zebrafish (Danio rerio) embryos to investigate the in vivo effects of FICZ when CYP1A is knocked down or inhibited. Embryos were injected with morpholino antisense oligonucleotides targeting CYP1A (CYP1A-MO), Ahr2, or a combination of both. FICZ exposure of non-injected embryos or embryos injected with control morpholino had little effect. In CYP1A-MO-injected embryos, however, FICZ dramatically increased mortality, incidence and severity of pericardial edema and circulation failure, reduced hatching frequency, blocked swim bladder inflation, and strongly potentiated expression of Ahr2-regulated genes. These effects were substantially reduced in embryos with a combined knockdown of Ahr2 and CYP1A, indicating that the toxicity was mediated at least partly by Ahr2. Co-exposure to the CYP1 inhibitor alpha-naphthoflavone (αNF) and FICZ had similar effects as the combination of CYP1A-MO and FICZ. HPLC analysis of FICZ-exposed embryos showed increased levels of FICZ after concomitant CYP1A-MO injection or αNF co-exposure. Together, these results show that a functioning CYP1/AHR feedback loop is crucial for regulation of AHR signaling by a potential physiological ligand in vivo and further highlights the role of CYP1 enzymes in regulating biological effects of FICZ.


Assuntos
Carbazóis/farmacologia , Citocromo P-450 CYP1A1/genética , Edema/genética , Regulação da Expressão Gênica no Desenvolvimento , Receptores de Hidrocarboneto Arílico/genética , Proteínas de Peixe-Zebra/genética , Animais , Benzoflavonas/farmacologia , Citocromo P-450 CYP1A1/antagonistas & inibidores , Citocromo P-450 CYP1A1/deficiência , Edema/induzido quimicamente , Edema/metabolismo , Edema/patologia , Embrião não Mamífero , Retroalimentação Fisiológica , Injeções , Morfolinos/genética , Morfolinos/metabolismo , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Pericárdio/efeitos dos fármacos , Pericárdio/metabolismo , Pericárdio/patologia , Receptores de Hidrocarboneto Arílico/agonistas , Receptores de Hidrocarboneto Arílico/antagonistas & inibidores , Receptores de Hidrocarboneto Arílico/metabolismo , Transdução de Sinais , Peixe-Zebra , Proteínas de Peixe-Zebra/agonistas , Proteínas de Peixe-Zebra/antagonistas & inibidores , Proteínas de Peixe-Zebra/metabolismo
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