RESUMEN
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.
Asunto(s)
Antineoplásicos/farmacología , Benzoflavonas/farmacología , Citocromo P-450 CYP1B1/antagonistas & inhibidores , Resistencia a Antineoplásicos/efectos de los fármacos , Neoplasias de la Próstata/tratamiento farmacológico , Antineoplásicos/síntesis química , Antineoplásicos/química , Benzoflavonas/síntesis química , Benzoflavonas/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Citocromo P-450 CYP1B1/metabolismo , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Masculino , Estructura Molecular , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Solubilidad , Relación Estructura-ActividadRESUMEN
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.
Asunto(s)
Benzoflavonas/farmacología , Citocromo P-450 CYP1B1/antagonistas & inhibidores , Resistencia a Antineoplásicos/efectos de los fármacos , Talidomida/análogos & derivados , Talidomida/farmacología , Benzoflavonas/síntesis química , Línea Celular Tumoral , Citocromo P-450 CYP1B1/metabolismo , Docetaxel/farmacología , Diseño de Fármacos , Humanos , Proteolisis , Talidomida/síntesis químicaRESUMEN
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.
Asunto(s)
Benzoflavonas/farmacología , Citocromo P-450 CYP1B1/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Benzoflavonas/síntesis química , Benzoflavonas/química , Citocromo P-450 CYP1B1/metabolismo , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Humanos , Simulación del Acoplamiento Molecular , Estructura Molecular , Proteínas Recombinantes/metabolismo , Relación Estructura-ActividadRESUMEN
Selective cytochrome P450 (CYP) 1B1 inhibition has potential as an anticancer strategy that is unrepresented in the current clinical arena. For development of a selective inhibitor, we focused on the complexity caused by sp3-hybridized carbons and synthesized a series of benzo[h]chromone derivatives linked to a non-aromatic B-ring using α-naphthoflavone (ANF) as the lead compound. Ring structure comparison suggested compound 37 as a suitable cyclohexyl-core with improved solubility. Structural evolution of 37 produced the azide-containing cis-49a, which had good properties in three important respects: (1) selectivity for CYP1B1 over CYP1A1 and CYP1A2 (120-times and 150-times, respectively), (2) greater inhibitory potency of >2 times that of ANF, and (3) improved solubility. The corresponding aromatic B-ring compound 59a showed low selectivity and poor solubility. To elucidate the binding mode, we performed X-ray crystal structure analysis, which revealed the interaction mode and explained the subtype selectivity of cis-49a.
Asunto(s)
Benzoflavonas/química , Inhibidores del Citocromo P-450 CYP1A2/química , Citocromo P-450 CYP1B1/antagonistas & inhibidores , Benzoflavonas/síntesis química , Dominio Catalítico , Cristalografía por Rayos X , Citocromo P-450 CYP1A1/antagonistas & inhibidores , Citocromo P-450 CYP1A1/química , Citocromo P-450 CYP1A2/química , Inhibidores del Citocromo P-450 CYP1A2/síntesis química , Citocromo P-450 CYP1B1/química , Diseño de Fármacos , Escherichia coli/genética , Humanos , Simulación del Acoplamiento Molecular , Estructura Molecular , Solubilidad , Relación Estructura-ActividadRESUMEN
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.
Asunto(s)
Benzoflavonas/farmacología , Inhibidores Enzimáticos/farmacología , Metiltestosterona/antagonistas & inhibidores , Oxigenasas de Función Mixta/antagonistas & inhibidores , beta-naftoflavona/farmacología , Absidia/enzimología , Benzoflavonas/síntesis química , Benzoflavonas/química , Chaetomium/enzimología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Metiltestosterona/química , Metiltestosterona/metabolismo , Oxigenasas de Función Mixta/metabolismo , Estructura Molecular , Mucorales/enzimología , Relación Estructura-Actividad , beta-naftoflavona/síntesis química , beta-naftoflavona/químicaRESUMEN
A library of forty 7,8-benzoflavone derivatives was synthesized and evaluated for their inhibitory potential against cholesterol esterase (CEase). Among all the synthesized compounds seven benzoflavone derivatives (A-7, A-8, A-10, A-11, A-12, A-13, A-15) exhibited significant inhibition against CEase in in vitro enzymatic assay. Compound A-12 showed the most promising activity with IC50 value of 0.78nM against cholesterol esterase. Enzyme kinetic studies carried out for A-12, revealed its mixed-type inhibition approach. Molecular protein-ligand docking studies were also performed to figure out the key binding interactions of A-12 with the amino acid residues of the enzyme's active site. The A-12 fits well at the catalytic site and is stabilized by hydrophobic interactions. It completely blocks the catalytic assembly of CEase and prevents it to participate in ester hydrolysis mechanism. The favorable binding conformation of A-12 suggests its prevailing role as CEase inhibitor.
Asunto(s)
Benzoflavonas/química , Inhibidores Enzimáticos/química , Esterol Esterasa/antagonistas & inhibidores , Benzoflavonas/síntesis química , Benzoflavonas/metabolismo , Sitios de Unión , Dominio Catalítico , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/metabolismo , Interacciones Hidrofóbicas e Hidrofílicas , Concentración 50 Inhibidora , Cinética , Simulación del Acoplamiento Molecular , Unión Proteica , Esterol Esterasa/metabolismo , Relación Estructura-ActividadRESUMEN
Multidrug resistance (MDR) often leads to a failure of cancer chemotherapy. Breast Cancer Resistance Protein (BCRP/ABCG2), a member of the superfamily of ATP binding cassette proteins has been found to confer MDR in cancer cells by transporting molecules with amphiphilic character out of the cells using energy from ATP hydrolysis. Inhibiting BCRP can be a solution to overcome MDR. We synthesized a series of flavones, 7,8-benzoflavones and 5,6-benzoflavones with varying substituents at positions 3, 3' and 4' of the (benzo)flavone structure. All synthesized compounds were tested for BCRP inhibition in Hoechst 33342 and pheophorbide A accumulation assays using MDCK cells expressing BCRP. All the compounds were further screened for their P-glycoprotein (P-gp) and Multidrug resistance-associated protein 1 (MRP1) inhibitory activity by calcein AM accumulation assay to check the selectivity towards BCRP. In addition most active compounds were investigated for their cytotoxicity. It was observed that in most cases 7,8-benzoflavones are more potent in comparison to the 5,6-benzoflavones. In general it was found that presence of a 3-OCH3 substituent leads to increase in activity in comparison to presence of OH or no substitution at position 3. Also, it was found that presence of 3',4'-OCH3 on phenyl ring lead to increase in activity as compared to other substituents. Compound 24, a 7,8-benzoflavone derivative was found to be most potent being 50 times selective for BCRP and showing very low cytotoxicity at higher concentrations.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/antagonistas & inhibidores , Benzoflavonas/farmacología , Flavonas/farmacología , Proteínas de Neoplasias/antagonistas & inhibidores , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Benzoflavonas/síntesis química , Benzoflavonas/química , Células Cultivadas , Perros , Relación Dosis-Respuesta a Droga , Flavonas/síntesis química , Flavonas/química , Humanos , Estructura Molecular , Proteínas de Neoplasias/metabolismo , Relación Estructura-ActividadRESUMEN
A phytotoxic root exudate from Acroptilon repens was identified as 7,8-benzoflavone, an inhibitor of cytochrome P450 1A2 and activator of cytochrome P450 3A4. The synthetic 5,6-benzoflavone also is a potent phytotoxin. Six 7,8-benzoflavones and eight 5,6-benzoflavones were synthesized in this study. The NMR data for a few of these compounds have been previously reported; however, the NMR data for most of them have not been reported. For reference purposes, the complete NMR data for the 14 benzoflavones are described.
Asunto(s)
Benzoflavonas/síntesis química , Benzoflavonas/química , Espectroscopía de Resonancia Magnética/normas , Estructura Molecular , Estándares de ReferenciaRESUMEN
Our previous screen of flavones and related heterocycles for the ability to activate the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel indicated that UCCF-029, a 7,8-benzoflavone, was a potent activator. In the present study, we describe the synthesis and evaluation, using cell-based assays, of a series of benzoflavone analogues to examine structure-activity relationships and to identify compounds having greater potency for activation of both wild type CFTR and a mutant CFTR (G551D-CFTR) that causes cystic fibrosis in some human subjects. Using UCCF-029 as a structural guide, a panel of 77 flavonoid analogues was prepared. Analysis of the panel in FRT cells indicated that benzannulation of the flavone A-ring at the 7,8-position greatly improved compound activity and potency for several flavonoids. Incorporation of a B-ring pyridyl nitrogen either at the 3- or 4-position also elevated CFTR activity, but the influence of this structural modification was not as uniform as the influence of benzannulation. The most potent new analogue, UCCF-339, activated wild-type CFTR with a K(d) of 1.7 microM, which is more active than the previous most potent flavonoid activator of CFTR, apigenin. Several compounds in the benzoflavone panel also activated G551D-CFTR, but none were as active as apigenin. Pharmacophore modeling suggests a common binding mode for the flavones and other known CFTR activators at one of the nucleotide-binding sites, allowing for the rational development of more potent flavone analogues.
Asunto(s)
Benzoflavonas/síntesis química , Regulador de Conductancia de Transmembrana de Fibrosis Quística/efectos de los fármacos , Nucleótidos/metabolismo , Animales , Apigenina , Benzoflavonas/farmacología , Sitios de Unión , Células Cultivadas , Regulador de Conductancia de Transmembrana de Fibrosis Quística/química , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Flavonoides/farmacología , Proteínas Fluorescentes Verdes , Humanos , Proteínas Luminiscentes , Modelos Moleculares , Conformación Molecular , Nucleótidos/química , Ratas , Relación Estructura-ActividadRESUMEN
A series of substituted and structural analogues of 7,8-benzoflavone were examined for their ability to inhibit benzo[a]pyrene oxidation by the mixed-function oxidases found in hepatic microsomes prepared from 3-methylcholanthrene- and phenobarbital-induced rats. Of all the benzoflavones tested, only 6-amino-7,8-benzoflavone possessed significant inhibitory activity toward both classes of induced mixed-function oxidases. Parameters which were found to be necessary for maximal inhibitory activity were the maintenance of an unsubstituted or specifically substituted exocyclic phenyl group on position 2, the preservation of the pyran-4-one ring, and a 6 position which is either unsubstituted or substituted with an oxidizable moiety.