RESUMEN
BACKGROUND: The phosphatidylinositol-3-kinase/mammalian target of rapamycin (PI3K/mTOR) pathway is commonly deregulated in human cancer, hence many PI3K and mTOR inhibitors have been developed and have now reached clinical trials. Similarly, CDKs have been investigated as cancer drug targets. METHODS: We have synthesised and characterised a series of 6-aminopyrimidines identified from a kinase screen that inhibit PI3K and/or mTOR and/or CDK2. Kinase inhibition, tumour cell growth, cell cycle distribution, cytotoxicity and signalling experiments were undertaken in HCT116 and HT29 colorectal cancer cell lines, and in vivo HT29 efficacy studies. RESULTS: 2,6-Diaminopyrimidines with an O(4)-cyclohexylmethyl substituent and a C-5-nitroso or cyano group (1,2,5) induced cell cycle phase alterations and were growth inhibitory (GI50<20 µM). Compound 1, but not 2 or 5, potently inhibits CDK2 (IC50=0.1 nM) as well as PI3K, and was cytotoxic at growth inhibitory concentrations. Consistent with kinase inhibition data, compound 1 reduced phospho-Rb and phospho-rS6 at GI50 concentrations. Combination of NU6102 (CDK2 inhibitor) and pictilisib (GDC-0941; pan-PI3K inhibitor) resulted in synergistic growth inhibition, and enhanced cytotoxicity in HT29 cells in vitro and HT29 tumour growth inhibition in vivo. CONCLUSIONS: These studies identified a novel series of mixed CDK2/PI3K inhibitors and demonstrate that dual targeting of CDK2 and PI3K can result in enhanced antitumour activity.
Asunto(s)
Adenocarcinoma/patología , Neoplasias Colorrectales/patología , Quinasa 2 Dependiente de la Ciclina/antagonistas & inhibidores , Terapia Molecular Dirigida , Proteínas de Neoplasias/antagonistas & inhibidores , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirimidinas/uso terapéutico , Adenocarcinoma/enzimología , Animales , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Neoplasias Colorrectales/enzimología , Quinasa 2 Dependiente de la Ciclina/fisiología , Humanos , Ratones , Ratones Desnudos , Proteínas de Neoplasias/fisiología , Fosfatidilinositol 3-Quinasas/fisiología , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/fisiología , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas/síntesis química , Pirimidinas/farmacología , Distribución Aleatoria , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Ensayo de Tumor de Célula Madre , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Combined targeting of the MAPK and PI3K signalling pathways in cancer may be necessary for optimal therapeutic activity. To support clinical studies of combination therapy, 3'-deoxy-3'-[(18)F]-fluorothymidine ([(18)F]-FLT) uptake measured by Positron Emission Tomography (PET) was evaluated as a non-invasive surrogate response biomarker in pre-clinical models. The in vivo anti-tumour efficacy and PK-PD properties of the MEK inhibitor PD 0325901 and the PI3K inhibitor GDC-0941, alone and in combination, were evaluated in HCT116 and HT29 human colorectal cancer xenograft tumour-bearing mice, and [(18)F]-FLT PET investigated in mice bearing HCT116 xenografts. Dual targeting of PI3K and MEK induced marked tumour growth inhibition in vivo, and enhanced anti-tumour activity was predicted by [(18)F]-FLT PET scanning after 2 days of treatment. Pharmacodynamic analyses using the combination of the PI3K inhibitor GDC-0941 and the MEK inhibitor PD 0325901 revealed that increased efficacy is associated with an enhanced inhibition of the phosphorylation of ERK1/2, S6 and 4EBP1, compared to that observed with either single agent, and maintained inhibition of AKT phosphorylation. Pharmacokinetic studies indicated that there was no marked PK interaction between the two drugs. Together these results indicate that the combination of PI3K and MEK inhibitors can result in significant efficacy, and demonstrate for the first time that [(18)F]-FLT PET can be correlated to the improved efficacy of combined PI3K and MEK inhibitor treatment.
Asunto(s)
Antivirales/farmacología , Benzamidas/farmacología , Neoplasias Colorrectales , Didesoxinucleósidos/farmacología , Difenilamina/análogos & derivados , Indazoles/farmacología , Quinasas Quinasa Quinasa PAM/antagonistas & inhibidores , Inhibidores de las Quinasa Fosfoinosítidos-3 , Tomografía de Emisión de Positrones , Sulfonamidas/farmacología , Animales , Benzamidas/agonistas , Línea Celular Tumoral , Neoplasias Colorrectales/diagnóstico por imagen , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/enzimología , Difenilamina/agonistas , Difenilamina/farmacología , Sinergismo Farmacológico , Femenino , Xenoinjertos , Humanos , Indazoles/agonistas , Quinasas Quinasa Quinasa PAM/metabolismo , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Radiografía , Sulfonamidas/agonistasRESUMEN
Amino-substituted biphenyls were obtained by Suzuki cross-coupling of 2,6-dibromoaniline with a phenylboronic acid (substituted with Me, NO(2), OH, OMe or Cl) preferably assisted by microwave irradiation. Conversion of the amino group into a thiol preceded a base-induced intramolecular substitution, also facilitated by microwave heating, to generate the second C-S bond of the target dibenzothiophene. The 1-, 2-, 3- or 4-substituted 6-halodibenzothiophenes obtained were subjected to a palladium-mediated coupling with 2-morpholin-4-yl-8-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4H-chromen-4-one to give the respective 6-, 7-, 8- or 9-substituted dibenzothiophen-4-ylchromenones. These compounds were evaluated as inhibitors of DNA-dependent protein kinase (DNA-PK) and compared to the parent 8-(dibenzo[b,d]thiophen-4-yl)-2-morpholin-4-yl-4H-chromen-4-one. Notably, derivatives bearing hydroxy or methoxy substituents at C-8 or C-9 retained activity, whereas substitution at C-7 lowered activity. Substitution with chloro at C-6 was not detrimental to activity, but a chloro group at C-7 or C-8 reduced potency. The data indicate permissive elaboration of hydroxyl at C-8 or C-9, enabling the possibility of improved pharmaceutical properties, whilst retaining potency against DNA-PK.
Asunto(s)
Proteína Quinasa Activada por ADN/antagonistas & inhibidores , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Tiofenos/síntesis química , Tiofenos/farmacología , Ciclización , Proteína Quinasa Activada por ADN/metabolismo , Inhibidores Enzimáticos/química , Humanos , Microondas , Tiofenos/químicaRESUMEN
Inhibition of the MDM2-p53 interaction has been shown to produce an antitumor effect, especially in MDM2 amplified tumors. The isoindolinone scaffold has proved to be versatile for the discovery of MDM2-p53 antagonists. Optimization of previously reported inhibitors, for example, NU8231 (7) and NU8165 (49), was guided by MDM2 NMR titrations, which indicated key areas of the binding interaction to be explored. Variation of the 2-N-benzyl and 3-alkoxy substituents resulted in the identification of 3-(4-chlorophenyl)-3-((1-(hydroxymethyl)cyclopropyl)methoxy)-2-(4-nitrobenzyl)isoindolin-1-one (74) as a potent MDM2-p53 inhibitor (IC(50) = 0.23 ± 0.01 µM). Resolution of the enantiomers of 74 showed that potent MDM2-p53 activity primarily resided with the (+)-R-enantiomer (74a; IC(50) = 0.17 ± 0.02 µM). The cellular activity of key compounds has been examined in cell lines with defined p53 and MDM2 status. Compound 74a activates p53, MDM2, and p21 transcription in MDM2 amplified cells and shows moderate selectivity for wild-type p53 cell lines in growth inhibition assays.
Asunto(s)
Antineoplásicos/síntesis química , Isoindoles/síntesis química , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Isoindoles/química , Isoindoles/farmacología , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Estructura Molecular , Unión Proteica , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
CDK2 inhibitory structure-activity relationships have been explored for a range of 5-substituted O(4)-alkylpyrimidines. Variation of the 5-substituent in the 2,6-diaminopyrimidine series confirmed the 5-nitroso substituent as optimal, and showed that 5-formyl and 5-acetyl substituents were also tolerated at this position. A series of O(4)-alkyl-N(2)-aryl-5-substituted-6-aminopyrimidines revealed interesting structure-activity relationships. In the 5-nitroso series, the optimum O(4)-alkyl substituents were cyclohexylmethyl or sec-butyl, combined with a 2-sulfanilyl group. By contrast, in the N(2)-arylsulfonamido-5-formyl series, the cyclohexylmethyl compound showed relatively poor activity compared with the sec-butyl derivative (22j, (R)-4-(4-amino-6-sec-butoxy-5-formylpyrimidin-2-ylamino)benzenesulfonamide; CDK2 IC(50) = 0.8 nM). Similarly, in the N(2)-arylsulfonamido-5-(hydroxyiminomethyl) series the O(4)-sec-butyl substituent conferred greater potency than the cyclohexylmethyl (23c, (rac)-4-(4-amino-6-sec-butoxy-5-(hydroxyiminomethyl)pyrimidin-2-ylamino)benzenesulfonamide; CDK2 IC(50) = 7.4 nM). The 5-formyl derivatives show selectivity for CDK2 over other CDK family members, and are growth inhibitory in tumour cells (e.g. 22j, GI(50) = 0.57 microM).
Asunto(s)
Quinasa 2 Dependiente de la Ciclina/antagonistas & inhibidores , Oxígeno/química , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas/síntesis química , Pirimidinas/farmacología , Línea Celular Tumoral , Descubrimiento de Drogas , Humanos , Concentración 50 Inhibidora , Inhibidores de Proteínas Quinasas/química , Pirimidinas/química , Relación Estructura-ActividadRESUMEN
Replacement of the core heterocycle of a defined series of chromen-4-one DNA-PK inhibitors by the isomeric chromen-2-one (coumarin) and isochromen-1-one (isocoumarin) scaffolds was investigated. Structure-activity relationships for DNA-PK inhibition were broadly consistent, albeit with a reduction of potency compared with the parent chromenone.