RESUMEN
The solution-phase, parallel synthesis and evaluation of a library of 132 (+)-1,2,9,9a-tetrahydrocyclopropa[c]benz[e]indol-4-one (CBI) analogues of CC-1065 and the duocarmycins containing dimeric monocyclic, bicyclic, and tricyclic heteroaromatic replacements for the DNA-binding domain are described. This systematic study revealed clear trends in the structural requirements for observation of potent cytotoxic activity and DNA alkylation efficiency, the range of which spans a magnitude of > or =10 000-fold. Combined with related studies, these results highlight that the role of the DNA-binding domain goes beyond simply providing DNA-binding selectivity and affinity (10-100-fold enhancement in properties), consistent with the proposal that it contributes significantly to catalysis of the DNA alkylation reaction accounting for as much as an additional 1000-fold enhancement in properties.
Asunto(s)
Antibióticos Antineoplásicos/síntesis química , Antibióticos Antineoplásicos/farmacología , Leucomicinas/síntesis química , Alquilantes/síntesis química , Alquilantes/química , Alquilantes/farmacología , Alquilación/efectos de los fármacos , Antibióticos Antineoplásicos/química , Sitios de Unión , Técnicas Químicas Combinatorias , Ciclopropanos/química , ADN/metabolismo , ADN Viral/efectos de los fármacos , ADN Viral/metabolismo , Duocarmicinas , Indoles/química , Concentración 50 Inhibidora , Leucomicinas/química , Leucomicinas/farmacología , Pirroles/síntesis química , Pirroles/química , Pirroles/farmacología , Pirrolidinonas/síntesis química , Pirrolidinonas/química , Pirrolidinonas/farmacología , Virus 40 de los Simios/genética , Relación Estructura-ActividadRESUMEN
While the endogenous fatty acid amide oleamide has hypnotic properties, neither the breadth of its behavioral actions nor the mechanism(s) by which these behaviors may be mediated has been elucidated. Therefore, the effects of oleamide on the performance of rats in tests of motor function, analgesia, and anxiety were investigated. Oleamide reduced the distance traveled in the open field (ED50 = 14, 10-19 mg/kg, mean, 95% confidence interval), induced analgesia and hypothermia, but did not cause catalepsy. Moreover, a dose of oleamide without effect on motor function was anxiolytic in the social interaction test and elevated plus-maze. These actions of a single dose of oleamide lasted for 30 to 60 min. While rats became tolerant to oleamide following 8 days of repeated administration, oleamide is a poor inducer of physical dependence. Pretreatment with antagonists of the serotonin (5HT)1A, 5HT2C, and vanilloid receptors did not modify oleamide's effects. However, the cannabinoid receptor antagonist SR 141716A inhibited oleamide-induced analgesia in the tail-flick assay, the gamma-aminobutyric acid (GABA)A receptor antagonist bicuculline reversed the analgesia and hypothermia, and the dopamine D2 receptor antagonist L 741626 blocked oleamide's locomotor and analgesic actions. Interestingly, oleamide analogs resistant to hydrolysis by fatty acid amide hydrolase (FAAH) maintained but did not show increased behavioral potency or duration of action, whereas two FAAH inhibitors produced analogous behavioral effects. Thus, oleamide induces behaviors reminiscent of the actions of endogenous cannabinoids, but the involvement of GABAergic and dopaminergic systems, either directly or indirectly, in the actions of oleamide cannot be ruled out.
Asunto(s)
Conducta Animal/efectos de los fármacos , Neurotransmisores/fisiología , Ácidos Oléicos/farmacología , Amidohidrolasas/metabolismo , Animales , Ansiedad/psicología , Temperatura Corporal/efectos de los fármacos , Catalepsia/inducido químicamente , Tolerancia a Medicamentos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Masculino , Actividad Motora/efectos de los fármacos , Ácidos Oléicos/efectos adversos , Ácidos Oléicos/síntesis química , Dimensión del Dolor/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Tiempo de Reacción/efectos de los fármacos , Conducta Social , Síndrome de Abstinencia a Sustancias/psicologíaRESUMEN
Full details of the total syntheses of thiocoraline (1) and BE-22179 (2), C(2) symmetric bicyclic octadepsipeptides possessing two pendant 3-hydroxyquinoline chromophores, are described in which their relative and absolute stereochemistry were established. Key elements of the approach include the late-stage introduction of the chromophore, symmetrical tetrapeptide coupling, macrocyclization of the 26-membered octadepsipeptide conducted at the single secondary amide site following disulfide formation, and a convergent assemblage of the tetradepsipeptide with introduction of the labile thiol ester linkage in the final coupling reaction under near racemization free conditions. By virtue of the late-stage introduction of the chromophore and despite the challenges this imposes on the synthesis, this approach provides ready access to a range of key chromophore analogues. Thiocoraline and BE-22179 were shown to bind to DNA by high-affinity bisintercalation analogous to echinomycin, but with little or no perceptible sequence selectivity. Both 1 and 2 were found to exhibit exceptional cytotoxic activity (IC(50) = 200 and 400 pM, respectively, L1210 cell line) comparable to echinomycin and one analogue, which bears the luzopeptin chromophore, was also found to be a potent cytotoxic agent.
Asunto(s)
Antibacterianos/síntesis química , ADN/metabolismo , Depsipéptidos , Péptidos , Animales , Antibacterianos/metabolismo , Antibacterianos/farmacología , Antibióticos Antineoplásicos/síntesis química , Antibióticos Antineoplásicos/metabolismo , Antibióticos Antineoplásicos/farmacología , Bovinos , Supervivencia Celular/efectos de los fármacos , Equinomicina/metabolismo , Equinomicina/farmacología , VIH-1/enzimología , Concentración 50 Inhibidora , Ratones , Inhibidores de la Transcriptasa Inversa , Células Tumorales CultivadasRESUMEN
A series of CBI analogues of the duocarmycins and CC-1065 exploring substituent effects within the first indole DNA binding subunit are detailed. Substitution at the indole C5 position led to cytotoxic potency enhancements that are > or =1000-fold, providing simplified analogues containing a single DNA binding subunit that are more potent (IC(50)=2-3 pM) than CBI-TMI, duocarmycin SA, or CC-1065.
Asunto(s)
Antibióticos Antineoplásicos/síntesis química , ADN/química , Leucomicinas/síntesis química , Pirroles/síntesis química , Pirrolidinonas/síntesis química , Adenina/análogos & derivados , Adenina/química , Adenina/metabolismo , Alquilación , Antibióticos Antineoplásicos/farmacología , Sitios de Unión/fisiología , ADN/metabolismo , Aductos de ADN/química , Aductos de ADN/metabolismo , Duocarmicinas , Humanos , Indoles/química , Concentración 50 Inhibidora , Leucomicinas/química , Leucomicinas/metabolismo , Leucomicinas/farmacología , Conformación de Ácido Nucleico , Pirroles/farmacología , Pirrolidinonas/farmacología , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
Two sets of novel analogues of the recently disclosed alpha-keto heterocycle inhibitors of fatty acid amide hydrolase (FAAH), the enzyme responsible for regulation of endogenous oleamide and anandamide, were synthesized and evaluated in order to clarify a role of the electrophilic carbonyl group and structural features important for their activity. Both the electrophilic carbonyl and the degree of alpha-substitution markedly affect inhibitor potency.
Asunto(s)
Amidohidrolasas/antagonistas & inhibidores , Compuestos Heterocíclicos con 2 Anillos/farmacología , Amidohidrolasas/metabolismo , Animales , Ácidos Araquidónicos/farmacología , Membrana Celular/enzimología , Endocannabinoides , Inhibidores Enzimáticos/farmacología , Cetonas/farmacología , Cinética , Hígado/enzimología , Hígado/ultraestructura , Ácidos Oléicos/farmacología , Alcamidas Poliinsaturadas , Ratas , Relación Estructura-ActividadRESUMEN
Full details of the development of a simple, nondestructive, and high-throughput method for establishing DNA binding affinity and sequence selectivity are described. The method is based on the loss of fluorescence derived from the displacement of ethidium bromide or thiazole orange from the DNA of interest or, in selected instances, the change in intrinsic fluorescence of a DNA binding agent itself and is applicable for assessing relative or absolute DNA binding affinities. Enlisting a library of hairpin deoxyoligonucleotides containing all five base pair (512 hairpins) or four base pair (136 hairpins) sequences displayed in a 96-well format, a compound's rank order binding to all possible sequences is generated, resulting in a high-resolution definition of its sequence selectivity using this fluorescent intercalator displacement (FID) assay. As such, the technique complements the use of footprinting or affinity cleavage for the establishment of DNA binding selectivity and provides the information at a higher resolution. The merged bar graphs generated by this rank order binding provide a qualitative way to compare, or profile, DNA binding affinity and selectivity. The 96-well format assay (512 hairpins) can be conducted at a minimal cost (presently ca. $100 for hairpin deoxyoligonucleotides/assay with ethiduim bromide or less with thiazole orange), with a rapid readout using a fluorescent plate reader (15 min), and is adaptable to automation (Tecan Genesis Workstation 100 robotic system). Its use in generating a profile of DNA binding selectivity for several agents including distamycin A, netropsin, DAPI, Hoechst 33258, and berenil is described. Techniques for establishing binding constants from quantitative titrations are compared, and recommendations are made for use of a Scatchard or curve fitting analysis of the titration binding curves as a reliable means to quantitate the binding affinity.
Asunto(s)
ADN/química , Oligodesoxirribonucleótidos/química , Emparejamiento Base , Secuencia de Bases , Sitios de Unión , Bisbenzimidazol/química , Distamicinas/química , Colorantes Fluorescentes , Indoles/química , Netropsina/química , Conformación de Ácido Nucleico , Robótica , Análisis de Secuencia de ADN/métodos , Relación Estructura-ActividadRESUMEN
The synthesis of 5-methoxycarbonyl-1,2,9,9a-tetrahydrocyclopropa[c]benz[e]indol-4-one (C5-CO2Me-CBI), a substituted CBI derivative bearing a C5 methoxycarbonyl group, and its corresponding 5-hydroxymethyl derivative are described in efforts to establish substituent electronic effects on the agents' functional reactivity and the resulting effect this has on their rate of DNA alkylation. Resolution of an immediate C5-CO2Me-CBI precursor and its incorporation into both enantiomers of 16 and 17, analogues of the duocarmycins, are also detailed. A study of the solvolysis reactivity and regioselectivity of N-BOC-C5-CO2Me-CBI (12) revealed that the introduction of a C5 methyl ester modestly slowed the rate of solvolysis (1.8x, pH 3) without altering the inherent reaction regioselectivity (>20:1). The comparison of the X-ray structures of the N-CO2Me derivatives of C5-CO2Me-CBI and CBI revealed correlations with the reaction regioselectivity and the relative reactivity of the compounds. The latter correlated well with the less reactive C5-CO2Me-CBI exhibiting a shortened N2-C2a bond length (1.386 vs 1.390 A) and smaller chi1 dihedral angle (8.1 degrees vs 21.2 degrees ) indicative of greater vinylogous amide conjugation and was accompanied by a diminished (cross-conjugated) cyclopropane conjugation (shorter bond lengths). Establishment of the DNA alkyation properties revealed that C5-CO2Me-CBI-based agents retained the identical alkylation selectivity of the natural products. More importantly, the C5 methyl ester was found to decrease the rate (0.77x) of DNA alkylation relative to CBI, consistent with its inherent lower reactivity. These results indicate that the previously observed increase in the rate of DNA alkylation for C7-substituted CBI analogues including CCBI (7-cyano-CBI) is contrary to expectations based on their inherent reactivities. Unlike 17, in which the C5 methyl ester does not bind in the minor groove, the C7 substituent lies in the minor groove extending the rigid length of the agents, further enhancing the DNA binding-induced conformational change responsible for activation toward nucleophilic attack and catalysis of the DNA alkylation reaction.
Asunto(s)
Antibióticos Antineoplásicos/síntesis química , Antibióticos Antineoplásicos/farmacología , Antineoplásicos Alquilantes/síntesis química , Antineoplásicos Alquilantes/farmacología , Indoles , Leucomicinas/síntesis química , Leucomicinas/farmacología , Pirrolidinonas/síntesis química , Pirrolidinonas/farmacología , Alquilación , Animales , Antibióticos Antineoplásicos/química , Antineoplásicos Alquilantes/química , Cristalografía por Rayos X , ADN/efectos de los fármacos , ADN/metabolismo , Duocarmicinas , Concentración 50 Inhibidora , Cinética , Leucomicinas/química , Leucemia L1210/tratamiento farmacológico , Ratones , Pirroles/síntesis química , Pirroles/química , Pirroles/farmacología , Pirrolidinonas/químicaRESUMEN
The solution-phase synthesis of two 1000-membered positional scanning libraries of distamycin A analogues is described enlisting acid/base liquid-liquid extractions for isolation and purification of all intermediates and final products. The results of their screening for functional activity (L1210 cytotoxic potency) and DNA binding affinity were compared with those derived from libraries containing the same compound members but prepared in a smaller 10-compound mixture format. The positional scanning libraries, which are substantially less demanding to prepare, allowed the accurate detection of the global observations and the clearly more potent activities, but more subtle discoveries and less distinguishable activities were not detected. This is a natural consequence of testing the larger 100-compound mixtures and the relative insensitivity of the assays to the contribution of any single, uniquely acting compound in the mixture. Thus, the disadvantages associated with the loss of some information contained within the library must be balanced against the advantages of the ease of library synthesis and judged in light of the library screening objectives.
Asunto(s)
Técnicas Químicas Combinatorias/métodos , ADN/metabolismo , Distamicinas/química , Distamicinas/metabolismo , Biblioteca de Péptidos , Animales , Diseño de Fármacos , Ensayos de Selección de Medicamentos Antitumorales , Leucemia L1210 , Ratones , Estructura MolecularRESUMEN
The examination results of a novel series of potential inhibitors of glycinamide ribonucleotide transformylase (GAR Tfase) and aminoimidazole carboxamide transformylase (AICAR Tfase) are reported. These agents incorporate an electrophilic fluoronitrophenyl group that can potentially react with an active site nucleophile or the substrate GAR/AICAR amine via nucleophilic aromatic substitution.
Asunto(s)
Inhibidores Enzimáticos/síntesis química , Ácido Fólico/análogos & derivados , Transferasas de Hidroximetilo y Formilo/antagonistas & inhibidores , Nitrocompuestos/síntesis química , Ribonucleótidos/metabolismo , Animales , Bioensayo , Técnicas Químicas Combinatorias , Diseño de Fármacos , Inhibidores Enzimáticos/farmacología , Flúor/química , Ácido Fólico/farmacología , Estructura Molecular , Fosforribosilglicinamida-Formiltransferasa , Leucemia-Linfoma Linfoblástico de Células Precursoras , Purinas/biosíntesis , Células Tumorales CultivadasRESUMEN
The synthesis and evaluation of a series of conformationally restricted analogues of 10-formyl-tetrahydrofolate as potential inhibitors of glycinamide ribonucleotide transformylase (GAR Tfase) or aminoimidazole carboxamide transformylase (AICAR Tfase) are reported.
Asunto(s)
Antagonistas del Ácido Fólico/química , Transferasas de Hidroximetilo y Formilo/antagonistas & inhibidores , Tetrahidrofolato Deshidrogenasa/efectos de los fármacos , Timidilato Sintasa/antagonistas & inhibidores , Línea Celular , Supervivencia Celular , Antagonistas del Ácido Fólico/farmacología , Humanos , Espectroscopía de Resonancia Magnética , Espectrometría de Masas , Estructura Molecular , Fosforribosilglicinamida-FormiltransferasaRESUMEN
The synthesis of 1,2,8,8a-tetrahydrocyclopropa[c]pyrrolo[3, 2-e]indol-4(5H)-one (CPI), the parent CC-1065 and duocarmycin SA alkylation subunit, is detailed. The parent CPI alkylation subunit lacks the C7 methyl substituent of the CC-1065 alkylation subunit and the C6 methoxycarbonyl group of duocarmycin SA, and their examination permitted the establishment of the impact of these natural product substituents. The studies revealed a CPI stability comparable to the CC-1065 alkylation subunit but which was 6x more reactive than the (+)-duocarmycin SA alkylation subunit, and it displayed the inherent reaction regioselectivity (4:1) of the natural products. The single-crystal X-ray structure of (+)-N-BOC-CPI depicts a near identical stereoelectronic alignment of the cyclopropane accounting for the identical reaction regioselectivity and a slightly diminished vinylogous amide conjugation relative to (+)-N-BOC-DSA suggesting that the stability distinctions stem in part from this difference in the vinylogous amide as well as alterations in the electronic nature of the fused pyrrole. Establishment of the DNA binding properties revealed that the CPI-based agents retain the identical DNA alkylation selectivities of the natural products. More importantly, the C6 methoxycarbonyl group of duocarmycin SA was found to increase the rate (12-13x) and efficiency (10x) of DNA alkylation despite its intrinsic lower reactivity while the CC-1065 C7 methyl group was found to slow the DNA alkylation rate (4x) and lower the alkylation efficiency (ca. 4x). The greater DNA alkylation rate and efficiency for duocarmycin SA and related analogues containing the C6 methoxycarbonyl is proposed to be derived from the extended length that the rigid C6 methoxycarbonyl provides and the resulting increase in the DNA binding-induced conformational change which serves to deconjugate the vinylogous amide and activate the alkylation subunit for nucleophilic attack. The diminished properties resulting from the CC-1065 C7 methyl group may be attributed to the steric impediment this substituent introduces to DNA minor groove binding and alkylation. Consistent with this behavior, the duocarmycin SA C6 methoxycarbonyl group increases biological potency while the CC-1065 C7 methyl group diminishes it.
Asunto(s)
Alquilantes/química , Antibióticos Antineoplásicos/química , ADN/química , Indolquinonas , Indoles , Leucomicinas/química , Quinolonas/síntesis química , Alquilación , Secuencia de Bases , Cristalografía por Rayos X , ADN Viral/química , Duocarmicinas , Indicadores y Reactivos , Modelos Moleculares , Conformación Molecular , Estructura Molecular , Oligodesoxirribonucleótidos/química , Pirroles/química , Quinolonas/químicaRESUMEN
The discovery of a novel class of diketopiperazines possessing potent cytotoxic activity is described.
Asunto(s)
ADN/metabolismo , Indoles/síntesis química , Indoles/farmacología , Piperazinas/síntesis química , Piperazinas/farmacología , Animales , Técnicas Químicas Combinatorias , Evaluación Preclínica de Medicamentos/métodos , Indoles/metabolismo , Concentración 50 Inhibidora , Leucemia L1210/tratamiento farmacológico , Piperazinas/metabolismo , Relación Estructura-ActividadRESUMEN
A concise, efficient approach to the total synthesis of ningalin B (1) based on a heterocyclic azadiene Diels-Alder strategy (1,2,4,5-tetrazine-->1,2,-diazine-->pyrrole) ideally suited for construction of the densely functionalized pyrrole core found in the natural product is detailed. Examination of the natural product and a number of synthetic intermediates revealed that while lacking inherent cytotoxic activity, many reverse the multidrug-resistant (MDR) phenotype, resensitizing a human colon cancer cell line (HCT116/VM46) to vinblastine and doxorubicin at lower doses than the prototypical agent verapamil.
Asunto(s)
Antineoplásicos/síntesis química , Compuestos Heterocíclicos con 3 Anillos/síntesis química , Urocordados/química , Animales , Antibióticos Antineoplásicos/farmacología , Supervivencia Celular/efectos de los fármacos , Doxorrubicina/farmacología , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Humanos , Ratones , Células Tumorales CultivadasRESUMEN
The development of exceptionally potent inhibitors of fatty acid amide hydrolase (FAAH), the enzyme responsible for the degradation of oleamide (an endogenous sleep-inducing lipid), and anandamide (an endogenous ligand for cannabinoid receptors) is detailed. The inhibitors may serve as useful tools to clarify the role of endogenous oleamide and anandamide and may prove to be useful therapeutic agents for the treatment of sleep disorders or pain. The combination of several features-an optimal C12-C8 chain length, pi-unsaturation introduction at the corresponding arachidonoyl Delta(8,9)/Delta(11,12) and oleoyl Delta(9,10) location, and an alpha-keto N4 oxazolopyridine with incorporation of a second weakly basic nitrogen provided FAAH inhibitors with K(i)s that drop below 200 pM and are 10(2)-10(3) times more potent than the corresponding trifluoromethyl ketones.