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1.
Nucleic Acids Res ; 52(4): 1736-1752, 2024 Feb 28.
Artículo en Inglés | MEDLINE | ID: mdl-38109306

RESUMEN

Repair of DNA damage is essential for the maintenance of genome stability and cell viability. DNA double strand breaks (DSBs) constitute a toxic class of DNA lesion and multiple cellular pathways exist to mediate their repair. Robust and titratable assays of cellular DSB repair (DSBR) are important to functionally interrogate the integrity and efficiency of these mechanisms in disease models as well as in response to genetic or pharmacological perturbations. Several variants of DSBR reporters are available, however these are often limited by throughput or restricted to specific cellular models. Here, we describe the generation and validation of a suite of extrachromosomal reporter assays that can efficiently measure the major DSBR pathways of homologous recombination (HR), classical nonhomologous end joining (cNHEJ), microhomology-mediated end joining (MMEJ) and single strand annealing (SSA). We demonstrate that these assays can be adapted to a high-throughput screening format and that they are sensitive to pharmacological modulation, thus providing mechanistic and quantitative insights into compound potency, selectivity, and on-target specificity. We propose that these reporter assays can serve as tools to dissect the interplay of DSBR pathway networks in cells and will have broad implications for studies of DSBR mechanisms in basic research and drug discovery.


Asunto(s)
Reparación del ADN , Ensayos Analíticos de Alto Rendimiento , ADN/metabolismo , Roturas del ADN de Doble Cadena , Reparación del ADN por Unión de Extremidades , Reparación del ADN/genética , Recombinación Homóloga , Reparación del ADN por Recombinación , Humanos , Línea Celular
3.
Clin Cancer Res ; 29(8): 1631-1642, 2023 04 14.
Artículo en Inglés | MEDLINE | ID: mdl-36689546

RESUMEN

PURPOSE: DNA polymerase theta (Polθ, encoded by the POLQ gene) is a DNA repair enzyme critical for microhomology mediated end joining (MMEJ). Polθ has limited expression in normal tissues but is frequently overexpressed in cancer cells and, therefore, represents an ideal target for tumor-specific radiosensitization. In this study we evaluate whether targeting Polθ with novel small-molecule inhibitors is a feasible strategy to improve the efficacy of radiotherapy. EXPERIMENTAL DESIGN: We characterized the response to Polθ inhibition in combination with ionizing radiation in different cancer cell models in vitro and in vivo. RESULTS: Here, we show that ART558 and ART899, two novel and specific allosteric inhibitors of the Polθ DNA polymerase domain, potently radiosensitize tumor cells, particularly when combined with fractionated radiation. Importantly, noncancerous cells were not radiosensitized by Polθ inhibition. Mechanistically, we show that the radiosensitization caused by Polθ inhibition is most effective in replicating cells and is due to impaired DNA damage repair. We also show that radiosensitization is still effective under hypoxia, suggesting that these inhibitors may help overcome hypoxia-induced radioresistance. In addition, we describe for the first time ART899 and characterize it as a potent and specific Polθ inhibitor with improved metabolic stability. In vivo, the combination of Polθ inhibition using ART899 with fractionated radiation is well tolerated and results in a significant reduction in tumor growth compared with radiation alone. CONCLUSIONS: These results pave the way for future clinical trials of Polθ inhibitors in combination with radiotherapy.


Asunto(s)
Neoplasias , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/radioterapia , Línea Celular Tumoral
4.
J Med Chem ; 65(24): 16589-16621, 2022 12 22.
Artículo en Inglés | MEDLINE | ID: mdl-36455032

RESUMEN

Small molecule inhibitors that target the phosphatidylinositol 3-kinase (PI3K) signaling pathway have received significant interest for the treatment of cancers. The class I isoform PI3Kα is most commonly associated with solid tumors via gene amplification or activating mutations. However, inhibitors demonstrating both PI3K isoform and mutant specificity have remained elusive. Herein, we describe the optimization and characterization of a series of benzoxazepin-oxazolidinone ATP-competitive inhibitors of PI3Kα which also induce the selective degradation of the mutant p110α protein, the catalytic subunit of PI3Kα. Structure-based design informed isoform-specific interactions within the binding site, leading to potent inhibitors with greater than 300-fold selectivity over the other Class I PI3K isoforms. Further optimization of pharmacokinetic properties led to excellent in vivo exposure and efficacy and the identification of clinical candidate GDC-0077 (inavolisib, 32), which is now under evaluation in a Phase III clinical trial as a treatment for patients with PIK3CA-mutant breast cancer.


Asunto(s)
Neoplasias de la Mama , Fosfatidilinositol 3-Quinasas , Humanos , Femenino , Inhibidores de las Quinasa Fosfoinosítidos-3/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3/uso terapéutico , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasa Clase I/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Línea Celular Tumoral , Mutación
5.
J Med Chem ; 65(20): 13879-13891, 2022 10 27.
Artículo en Inglés | MEDLINE | ID: mdl-36200480

RESUMEN

Human DNA polymerase theta (Polθ), which is essential for microhomology-mediated DNA double strand break repair, has been proposed as an attractive target for the treatment of BRCA deficient and other DNA repair pathway defective cancers. As previously reported, we recently identified the first selective small molecule Polθ in vitro probe, 22 (ART558), which recapitulates the phenotype of Polθ loss, and in vivo probe, 43 (ART812), which is efficacious in a model of PARP inhibitor resistant TNBC in vivo. Here we describe the discovery, biochemical and biophysical characterization of these probes including small molecule ligand co-crystal structures with Polθ. The crystallographic data provides a basis for understanding the unique mechanism of inhibition of these compounds which is dependent on stabilization of a "closed" enzyme conformation. Additionally, the structural biology platform provided a basis for rational optimization based primarily on reduced ligand conformational flexibility.


Asunto(s)
Reparación del ADN por Unión de Extremidades , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Humanos , Ligandos , ADN/metabolismo , ADN Polimerasa theta
6.
Nat Commun ; 12(1): 3636, 2021 06 17.
Artículo en Inglés | MEDLINE | ID: mdl-34140467

RESUMEN

To identify approaches to target DNA repair vulnerabilities in cancer, we discovered nanomolar potent, selective, low molecular weight (MW), allosteric inhibitors of the polymerase function of DNA polymerase Polθ, including ART558. ART558 inhibits the major Polθ-mediated DNA repair process, Theta-Mediated End Joining, without targeting Non-Homologous End Joining. In addition, ART558 elicits DNA damage and synthetic lethality in BRCA1- or BRCA2-mutant tumour cells and enhances the effects of a PARP inhibitor. Genetic perturbation screening revealed that defects in the 53BP1/Shieldin complex, which cause PARP inhibitor resistance, result in in vitro and in vivo sensitivity to small molecule Polθ polymerase inhibitors. Mechanistically, ART558 increases biomarkers of single-stranded DNA and synthetic lethality in 53BP1-defective cells whilst the inhibition of DNA nucleases that promote end-resection reversed these effects, implicating these in the synthetic lethal mechanism-of-action. Taken together, these observations describe a drug class that elicits BRCA-gene synthetic lethality and PARP inhibitor synergy, as well as targeting a biomarker-defined mechanism of PARPi-resistance.


Asunto(s)
Proteína BRCA1/genética , Proteína BRCA2/genética , Reparación del ADN/efectos de los fármacos , ADN Polimerasa Dirigida por ADN/genética , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Mutaciones Letales Sintéticas/efectos de los fármacos , Regulación Alostérica , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Proteína BRCA1/metabolismo , Proteína BRCA2/metabolismo , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Daño del ADN/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , ADN Polimerasa Dirigida por ADN/metabolismo , Desoxirribonucleasas/antagonistas & inhibidores , Resistencia a Antineoplásicos , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Recombinación Homóloga/efectos de los fármacos , Humanos , Concentración 50 Inhibidora , Ratones , Organoides/efectos de los fármacos , Neoplasias Ováricas/genética , Ratas , Mutaciones Letales Sintéticas/genética , Proteína 1 de Unión al Supresor Tumoral P53/deficiencia , Proteína 1 de Unión al Supresor Tumoral P53/metabolismo , ADN Polimerasa theta
7.
ACS Med Chem Lett ; 8(9): 936-940, 2017 Sep 14.
Artículo en Inglés | MEDLINE | ID: mdl-28947940

RESUMEN

A novel selective benzoxazepin inhibitor of PI3Kδ has been discovered. Beginning from compound 3, an αPI3K inhibitor, we utilized structure-based drug design and computational analysis of dihedral torsion angles to optimize for PI3Kδ isoform potency and isoform selectivity. Further medicinal chemistry optimization of the series led to the identification of 24, a highly potent and selective inhibitor of PI3Kδ.

8.
J Med Chem ; 59(19): 9080-9093, 2016 10 13.
Artículo en Inglés | MEDLINE | ID: mdl-27564586

RESUMEN

Inhibitors targeting the activating mutants of the epidermal growth factor receptor (EGFR) have found success in the treatment of EGFR mutant positive non-small-cell lung cancer. A secondary point mutation (T790M) in the inhibitor binding site has been linked to the acquired resistance against those first generation therapeutics. Herein, we describe the lead optimization of a series of reversible, pan-mutant (L858R, del746-750, T790M/L858R, and T790M/del746-750) EGFR inhibitors. By use of a noncovalent double mutant (T790M/L858R and T790M/del746-750) selective EGFR inhibitor (2) as a starting point, activities against the single mutants (L858R and del746-750) were introduced through a series of structure-guided modifications. The in vitro ADME-PK properties of the lead molecules were further optimized through a number of rational structural changes. The resulting inhibitor (21) exhibited excellent cellular activity against both the single and double mutants of EGFR, demonstrating target engagement in vivo and ADME-PK properties that are suitable for further evaluation. The reversible, noncovalent inhibitors described complement the covalent pan-mutant EGFR inhibitors that have shown encouraging results in recent clinical trials.


Asunto(s)
Antineoplásicos/química , Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Receptores ErbB/antagonistas & inhibidores , Neoplasias Pulmonares/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/uso terapéutico , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Resistencia a Antineoplásicos , Receptores ErbB/genética , Humanos , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Pulmón/patología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Ratones , Modelos Moleculares , Mutación , Inhibidores de Proteínas Quinasas/farmacocinética , Inhibidores de Proteínas Quinasas/farmacología
9.
J Med Chem ; 59(3): 985-1002, 2016 Feb 11.
Artículo en Inglés | MEDLINE | ID: mdl-26741947

RESUMEN

Inhibitors of the class I phosphoinositide 3-kinase (PI3K) isoform PI3Kα have received substantial attention for their potential use in cancer therapy. Despite the particular attraction of targeting PI3Kα, achieving selectivity for the inhibition of this isoform has proved challenging. Herein we report the discovery of inhibitors of PI3Kα that have selectivity over the other class I isoforms and all other kinases tested. In GDC-0032 (3, taselisib), we previously minimized inhibition of PI3Kß relative to the other class I insoforms. Subsequently, we extended our efforts to identify PI3Kα-specific inhibitors using PI3Kα crystal structures to inform the design of benzoxazepin inhibitors with selectivity for PI3Kα through interactions with a nonconserved residue. Several molecules selective for PI3Kα relative to the other class I isoforms, as well as other kinases, were identified. Optimization of properties related to drug metabolism then culminated in the identification of the clinical candidate GDC-0326 (4).


Asunto(s)
Antineoplásicos/farmacología , Benzoxepinas/farmacología , Diseño de Fármacos , Imidazoles/farmacología , Oxazepinas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/metabolismo , Benzoxepinas/química , Benzoxepinas/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Fosfatidilinositol 3-Quinasa Clase I , Perros , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Imidazoles/química , Imidazoles/metabolismo , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Macaca fascicularis , Ratones , Microsomas Hepáticos/química , Microsomas Hepáticos/metabolismo , Modelos Moleculares , Estructura Molecular , Oxazepinas/química , Oxazepinas/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/metabolismo , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad
10.
J Med Chem ; 57(23): 10176-91, 2014 Dec 11.
Artículo en Inglés | MEDLINE | ID: mdl-25383627

RESUMEN

Activating mutations within the epidermal growth factor receptor (EGFR) kinase domain, commonly L858R or deletions within exon 19, increase EGFR-driven cell proliferation and survival and are correlated with impressive responses to the EGFR inhibitors erlotinib and gefitinib in nonsmall cell lung cancer patients. Approximately 60% of acquired resistance to these agents is driven by a single secondary mutation within the EGFR kinase domain, specifically substitution of the gatekeeper residue threonine-790 with methionine (T790M). Due to dose-limiting toxicities associated with inhibition of wild-type EGFR (wtEGFR), we sought inhibitors of T790M-containing EGFR mutants with selectivity over wtEGFR. We describe the evolution of HTS hits derived from Jak2/Tyk2 inhibitors into selective EGFR inhibitors. X-ray crystal structures revealed two distinct binding modes and enabled the design of a selective series of novel diaminopyrimidine-based inhibitors with good potency against T790M-containing mutants of EGFR, high selectivity over wtEGFR, broad kinase selectivity, and desirable physicochemical properties.


Asunto(s)
Aminopiridinas/síntesis química , Resistencia a Antineoplásicos , Receptores ErbB/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/síntesis química , Sustitución de Aminoácidos , Aminopiridinas/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Cristalografía por Rayos X , Receptores ErbB/genética , Ensayos Analíticos de Alto Rendimiento , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Metionina/genética , Mutación , Inhibidores de Proteínas Quinasas/uso terapéutico , Treonina/genética
11.
J Med Chem ; 56(11): 4597-610, 2013 Jun 13.
Artículo en Inglés | MEDLINE | ID: mdl-23662903

RESUMEN

Dysfunctional signaling through the phosphoinositide 3-kinase (PI3K)/AKT/mTOR pathway leads to uncontrolled tumor proliferation. In the course of the discovery of novel benzoxepin PI3K inhibitors, we observed a strong dependency of in vivo antitumor activity on the free-drug exposure. By lowering the intrinsic clearance, we derived a set of imidazobenzoxazepin compounds that showed improved unbound drug exposure and effectively suppressed growth of tumors in a mouse xenograft model at low drug dose levels. One of these compounds, GDC-0032 (11l), was progressed to clinical trials and is currently under phase I evaluation as a potential treatment for human malignancies.


Asunto(s)
Antineoplásicos/síntesis química , Imidazoles/síntesis química , Oxazepinas/síntesis química , Inhibidores de las Quinasa Fosfoinosítidos-3 , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Hepatocitos/metabolismo , Humanos , Imidazoles/farmacocinética , Imidazoles/farmacología , Técnicas In Vitro , Isoenzimas/antagonistas & inhibidores , Ratones , Ratones Desnudos , Microsomas Hepáticos/metabolismo , Trasplante de Neoplasias , Oxazepinas/farmacocinética , Oxazepinas/farmacología , Relación Estructura-Actividad , Trasplante Heterólogo
12.
J Med Chem ; 55(10): 4594-604, 2012 May 24.
Artículo en Inglés | MEDLINE | ID: mdl-22506516

RESUMEN

Using structure-based design, two novel series of highly potent biaryl amine mitogen-activated protein kinase kinase (MEK) inhibitors have been discovered. These series contain an H-bond acceptor, in a shifted position compared with previously disclosed compounds, and an adjacent H-bond donor, resulting in a bidentate interaction with the Ser212 residue of MEK1. The most potent compound identified, 1 (G-894), is orally active in in vivo pharmacodynamic and tumor xenograft models.


Asunto(s)
Antineoplásicos/síntesis química , Benzofuranos/síntesis química , Benzotiazoles/síntesis química , Indazoles/síntesis química , MAP Quinasa Quinasa 1/antagonistas & inhibidores , MAP Quinasa Quinasa 2/antagonistas & inhibidores , Serina/metabolismo , Regulación Alostérica , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Benzofuranos/farmacocinética , Benzofuranos/farmacología , Benzotiazoles/farmacocinética , Benzotiazoles/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Enlace de Hidrógeno , Indazoles/farmacocinética , Indazoles/farmacología , Ratones , Modelos Moleculares , Estructura Molecular , Trasplante de Neoplasias , Ratas , Relación Estructura-Actividad , Trasplante Heterólogo
14.
J Med Chem ; 51(4): 963-75, 2008 Feb 28.
Artículo en Inglés | MEDLINE | ID: mdl-18247546

RESUMEN

The growth-inhibitory activities of an extensive series of quaternized quino[4,3,2- kl]acridinium salts against tumor cell lines in vitro have been measured and their biological properties interpreted in the light of differential binding to different DNA isoforms. Selectivity for quadruplex DNA binding and stabilization by compounds were explored through an array of methods: UV absorption and fluorescence emission spectroscopy, surface plasmon resonance, and competition dialysis. Quadruplex DNA interaction was further characterized through FRET and DNA polymerase arrest assays. Telomerase inhibition, inferred from the TRAP assay, is attributed to quadruplex stabilization, supported by the strong correlation (R(2) = 0.81) across the series between quadruplex DNA binding affinity and TRAP inhibition potency. Growth inhibition potency in the NCI60 human tumor cell line panel is more marked in compounds with greater DNA duplex binding affinity (R(2) = 0.82). Quantification of relative quadruplex and duplex binding affinity constants puts some of these ligands among the most selective quadruplex DNA interactive agents reported to date.


Asunto(s)
Acridinas/síntesis química , Antineoplásicos/síntesis química , G-Cuádruplex , Compuestos Heterocíclicos de 4 o más Anillos/síntesis química , Compuestos de Amonio Cuaternario/síntesis química , Telómero/metabolismo , Acridinas/química , Acridinas/farmacología , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , ADN/química , Ensayos de Selección de Medicamentos Antitumorales , Transferencia Resonante de Energía de Fluorescencia , Compuestos Heterocíclicos de 4 o más Anillos/química , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Humanos , Compuestos de Amonio Cuaternario/química , Compuestos de Amonio Cuaternario/farmacología , Relación Estructura-Actividad , Resonancia por Plasmón de Superficie , Telomerasa/antagonistas & inhibidores
15.
Bioorg Med Chem Lett ; 17(2): 363-9, 2007 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-17107790

RESUMEN

Further investigation of a series of thienyl-based hydroxamic acids that included ADS100380 and ADS102550 led to the identification of the 5-pyridin-2-yl-thiophene-2-hydroxamic acid 3c, which possessed modest HDAC inhibitory activity. Substitution at the 5- and 6-positions of the pyridyl ring of compound 3c provided compounds 5a-g, 7a, b, 9, and 13a. Compound 5b demonstrated improved potency, in vitro DMPK profile, and rat oral bioavailability, compared to ADS102550. Functionalisation of the pendent phenyl group of compounds 5b, 5e and 13a provided analogues that possessed excellent enzyme inhibition and anti-proliferative activity.


Asunto(s)
Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Inhibidores de Histona Desacetilasas , Ácidos Hidroxámicos/síntesis química , Ácidos Hidroxámicos/farmacología , Tiofenos/síntesis química , Tiofenos/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Disponibilidad Biológica , Células CACO-2 , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Citocromo P-450 CYP3A , Inhibidores Enzimáticos del Citocromo P-450 , Inhibidores Enzimáticos/farmacocinética , Humanos , Indicadores y Reactivos , Inyecciones Intravenosas , Ratones , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo , Ratas , Relación Estructura-Actividad
16.
J Med Chem ; 48(23): 7198-207, 2005 Nov 17.
Artículo en Inglés | MEDLINE | ID: mdl-16279778

RESUMEN

Palladium(0)-mediated Suzuki-Miyaura and Heck transformations have been exploited to provide examples of 8-methylquino[4,3,2-kl]acridines and 8,13-dimethylquino[4,3,2-kl]acridinium iodides bearing bulky saturated (3-acetoxy)propyl or (E)-3-(morpholin-4-yl)-3-oxopropenyl substituents variously in the 3-, 6-, or 10-positions of the pentacyclic nucleus. The pharmacological/pharmaceutical properties of four compounds (4, RHPS4), (5, IH383), (6, RHPS16), and (17, RHPS19) were measured to assess their clinical potential as DNA G-quadruplex-stabilizing/telomerase inhibitory agents. The following properties were measured: stability in tissue culture media in the presence of A549 lung and MCF-7 breast tumor cells, metabolic stability when incubated with rat liver microsomes, and rate of uptake and subcellular location in A549 and MCF-7 cells. Compound 17 was unstable in tissue culture media, failed to achieve nuclear access, and was excluded from further consideration. Of the other agents, 4 exhibited the most favorable pharmaceutical profile: the agent has appropriate stability in the presence of tumor cells and rat liver microsomes and achieves rapid ingress into cell nuclei where the putative molecular target is located.


Asunto(s)
Acridinas/síntesis química , Antineoplásicos/síntesis química , Compuestos Heterocíclicos de 4 o más Anillos/síntesis química , Telomerasa/antagonistas & inhibidores , Acridinas/química , Acridinas/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular Tumoral , Sistema Enzimático del Citocromo P-450/metabolismo , Ensayos de Selección de Medicamentos Antitumorales , Estabilidad de Medicamentos , Citometría de Flujo , Compuestos Heterocíclicos de 4 o más Anillos/química , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Humanos , Técnicas In Vitro , Microscopía Confocal , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/metabolismo , Ratas , Estereoisomerismo , Relación Estructura-Actividad
17.
Mol Pharmacol ; 68(6): 1551-8, 2005 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-16150933

RESUMEN

Telomeric integrity is required to maintain the replicative ability of cancer cells and is a target for the G-quadruplex-stabilizing drug 3,11-difluoro-6,8,13-trimethyl-8H-quino[4,3,2-kl]acridinium methosulfate (RHPS4). We report a senescent-like growth arrest in MCF-7 breast cancer cells, within 14 to 17 days, and a reduction in telomere length (from 5.2 kilobases (kb) to 4.7 and 4.3 kb after 17 days of treatment at 0.5 and 1 microM, respectively). These effects occurred at noncytotoxic drug concentrations (doses < 1 microM over a 14-day exposure) compatible with long-term drug dosing. The telomere length of cancer cells influences their sensitivity to growth inhibition by RHPS4: mutant (mt) human telomerase reverse transcriptase (hTERT)-expressing MCF-7 cells [short telomere restriction fragment (TRF) length, 1.9 kb; IC50, 0.2 microM] were 10 times more sensitive to RHPS4 compared with wild-type (wt) hTERT-expressing, vector-transfected control cells (longer TRF-length 5.2 kb; IC50 2 microM) in the 5 day SRB assay. This relationship was corroborated in a panel of 36 human tumor xenografts grown in vitro showing a positive correlation between telomere length and growth inhibitory potency of RHPS4 (15-day clonogenic assay, r = 0.75). These observations are consistent with loss of the protective capping status of telomeres mediated by RHPS4 G-quadruplex-stabilization, thus leading to greater susceptibility of cells with shorter telomeres. In combination studies, paclitaxel (Taxol), doxorubicin (Adriamycin), and the experimental therapeutic agent 17-(allylamino)-17-demethoxygeldanamycin, which inhibits the 90-kDa heat shock protein, conferred enhanced sensitivity in RHPS4 treated MCF-7 cells, whereas the DNA-interactive temozolomide and cisplatin antagonized the action of RHPS4. Our results support the combined use of certain classes of cytotoxic anticancer agents with RHPS4 to enhance potential clinical benefit.


Asunto(s)
Acridinas/farmacología , Antineoplásicos/farmacología , Proteínas de Unión al ADN/antagonistas & inhibidores , Telomerasa/antagonistas & inhibidores , Telómero/ultraestructura , Acridinas/farmacocinética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , ADN , Interacciones Farmacológicas , Inhibidores Enzimáticos/farmacología , G-Cuádruplex , Humanos , Farmacocinética
18.
J Med Chem ; 48(8): 2993-3004, 2005 Apr 21.
Artículo en Inglés | MEDLINE | ID: mdl-15828838

RESUMEN

The antileukemic xanthone psorospermin is a topoisomerase II-dependent DNA alkylator in advanced preclinical development. Efforts have been made to further understand the structural requirements of its mechanism of action through the synthesis of ring-constrained analogues, based on the skeleton of the bisfuranoxanthone natural products. Molecules were designed that contain the bisfuran and xanthone portions of naturally occurring psorofebrins, and molecular modeling was used to assess their DNA alkylating potential and to refine the structures. A short, diastereoselective synthetic process to access bisfuranoxanthones was developed, culminating in the first total synthesis of (+/-)-isohydroxypsorofebrin. Two compounds designed and synthesized were of particular interest, chlorohydrin 7 and epoxide 6, which are reactive analogues of the natural product isohydroxypsorofebrin. The chlorohydrin retains the psorospermin-like DNA alkylation characteristics despite its rigid structure and high innate affinity for DNA. Molecular modeling has been used to rationalize the increased activity of the chlorohydrin. The chlorohydrin and epoxide show increased cytotoxicity compared to isohydroxypsorofebrin against a range of human tumor cell lines.


Asunto(s)
Antineoplásicos Alquilantes/síntesis química , Furanos/síntesis química , Xantonas/síntesis química , Antineoplásicos Alquilantes/química , Antineoplásicos Alquilantes/farmacología , Línea Celular Tumoral , ADN-Topoisomerasas de Tipo II/metabolismo , Diseño de Fármacos , Ensayos de Selección de Medicamentos Antitumorales , Furanos/química , Furanos/farmacología , Humanos , Modelos Moleculares , Conformación Molecular , Estereoisomerismo , Relación Estructura-Actividad , Xantonas/química , Xantonas/farmacología
19.
Org Biomol Chem ; 2(2): 220-8, 2004 Jan 21.
Artículo en Inglés | MEDLINE | ID: mdl-14737646

RESUMEN

Cyclisation of 9-(benzotriazol-1-yl)acridine to the pentacycle 8H-quino[4,3,2-kl]acridine in a range of low-boiling solvents is mechanistically distinct from previously published photochemical (carbene) and thermolytic (radical) cyclisations. Fragmentation of the triazole ring of to a diazonium intermediate, and its subsequent heterolysis (-N(2)) and cyclisation is facilitated by solvation of intermediate zwitterionic species. Derivatives of 2- and 3-aminoquinoacridines methylated in the 8-position can be converted to 8,13-dimethylquino[4,3,2-kl]acridinium iodide salts with methyl iodide and were required for biological examination as potential telomerase inhibitors. The chloro group in 3-chloro-8-methyl-8H-quino[4,3,2-kl]acridine can be replaced efficiently by benzylamino, 4-morpholinyl and cyano substituents in palladium(0) mediated reactions.


Asunto(s)
Acridinas/síntesis química , Acridinas/farmacología , Acridinas/química , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Línea Celular Tumoral , Ciclización , Calor , Humanos , Concentración 50 Inhibidora , Solventes/química , Relación Estructura-Actividad , Triazoles/química
20.
J Mol Biol ; 334(1): 25-36, 2003 Nov 14.
Artículo en Inglés | MEDLINE | ID: mdl-14596797

RESUMEN

The NMR structure of the parallel-stranded DNA quadruplex d(TTAGGGT)(4), containing the human telomeric repeat, has been determined in solution in complex with a fluorinated pentacyclic quino[4,3,2-kl]acridinium cation (RHPS4). RHPS4 has been identified as a potent inhibitor of telomerase at submicromolar levels (IC(50) value of 0.33(+/-0.13)microM), exhibiting a wide differential between telomerase inhibition and acute cellular toxicity. All of the data point to RHPS4 exerting its chemotherapeutic potency through interaction with, and stabilisation of, four-stranded G-quadruplex structures. RHPS4 forms a dynamic interaction with d(TTAGGGT)(4), as evident from 1H and 19F linewidths, with fast exchange between binding sites induced at 318 K. Perturbations to DNA chemical shifts and 24 intermolecular nuclear Overhauser effects (NOEs) identify the 5'-ApG and 5'-GpT steps as the principle intercalation sites; a structural model has been refined using NOE-restrained molecular dynamics. The central G-tetrad core remains intact, with drug molecules stacking at the ends of the G-quadruplex. The partial positive charge on position 13-N of the acridine ring appears to act as a "pseudo" potassium ion and is positioned above the centre of the G-tetrad in the region of high negative charge density. In both ApG and GpT intercalation sites, the drug is seen to converge to the same orientation in which the pi-system of the drug overlaps primarily with two bases of each G-tetrad. The drug is held in place by stacking interactions with the G-tetrads; however, there is some evidence for a more dynamic, weakly stabilised A-tetrad that stacks partially on top of the drug at the 5'-end of the sequence. Together, the interactions of RHPS4 increase the t(m) of the quadruplex by approximately 20 degrees C. There is no evidence for drug intercalation within the G-quadruplex; however, the structural model strongly supports end-stacking interactions with the terminal G-tetrads.


Asunto(s)
Acridinas/química , ADN/química , Conformación de Ácido Nucleico , Telómero/genética , Acridinas/metabolismo , ADN/metabolismo , G-Cuádruplex , Humanos , Modelos Moleculares , Estructura Molecular , Resonancia Magnética Nuclear Biomolecular , Telomerasa/antagonistas & inhibidores
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