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J M Cook

Publications and source records attributed to J M Cook.

At least 55 records · Page 3Linked to original sources
PubMed1994

Synthesis of benzo-fused benzodiazepines employed as probes of the agonist pharmacophore of benzodiazepine receptors.

The synthesis and in vitro evaluation of benzo-fused benzodiazepines 1-6 are described. These "molecular yardsticks" were employed to probe the spatial dimensions of the lipophilic pocket L2 in the benzodiazepine receptor (BzR) cleft and to determine the effect of occupation of L2 with respect to agonist activity. Of the new analogs synthesized, the 7,8-benzo-fused benzodiazepine 6 displayed moderately high affinity for the BzR (IC50 = 55 nM) and exhibited both anticonvulsant (ED50 approximately 15 mg/kg) and muscle relaxant (ED50 approximately 15 mg/kg) activity. As expected, 2 and 4 interacted with the repulsive regions of interaction, S1 and S2, and exhibited low affinities for BzR. The rigid nature of these molecular yardsticks (especially 6, Figure 7) has been employed to probe the depth of L2. Moreover, in the case of 6 full occupation of L2 has resulted in an increase in the muscle relaxant effect at the expense of the anticonvulsant/anxiolytic effect.

Animals↗
PubMed1994

Epoxide metabolite of quinine and inhibition of the multidrug resistance pump in human leukemic lymphoblasts.

Multidrug resistance (MDR) in neoplastic cells is usually due to decreased cellular retention of drugs such as vincristine or doxorubicin. An ATP-dependent drug efflux pump has been detected in MDR-1-phenotypic cells; inhibition of the MDR pump is probably the primary mechanism for reversal of MDR. Although quinine (SQ1) and quinidine are reversal agents and inhibitors of the MDR pump, the results from in vivo experiments and in vitro experiments with these diastereomers are contradictory. These observations suggest that an oxidized metabolite of SQ1 is a more potent inhibitor of the MDR pump than is the parent compound. The chemical synthesis of the epoxides of SQ1 and quinidine is reported. The epoxy compounds have been tested as inhibitors of the ATP-dependent MDR pump in human CEM/VLB100 cells. The procedure is based on preloading the cells with an inhibitor and a low concentration of a substrate, rhodamine 123 (R123). After several cold rinses, the cell suspension is passed through a filtration-flow apparatus and the R123 in the filtrate (determined by fluorescence measurements) reveals the initial efflux of R123 through the MDR pump. When tested as an inhibitor of the MDR pump, quinine-10,11-epoxide is approximately 8-fold more potent than SQ1.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1993

Synthetic and computer-assisted analysis of the structural requirements for selective, high-affinity ligand binding to diazepam-insensitive benzodiazepine receptors.

Several 1,4-diazepines were recently reported to bind with high affinities to the "diazepam-insensitive" (DI) isoform of the benzodiazepine receptor (BzR) (Korpi, E.R.; Uusi-Oukari, M.; Wegelius, K. Eur. J. Pharm. 1992, 213, 323-329. Wong, G.; Skolnick, P. Eur. J. Pharmacol. Mol. Pharm. Sec. 1992, 225, 63-68). However, only the putative ethanol antagonist 1 (Ro 15-4513) displayed modest selectivity for the DI site compared to other "diazepam-sensitive" (DS) BzR isoforms. In order to probe the requirements for selective, high-affinity binding to the DI site, the affinities of 47 benzodiazepines have been determined at both DI and DS BzR sites. In addition, single X-ray crystallographic analyses for three of these derivatives, 5 (Ro 17-1812), 6 (Ro 16-6028), and 42 (Ro 14-5974), are reported. The radioligand binding studies reveal that modifications to the 3-, 7-, and 8-positions of 6-oxoimidazo[1,5-alpha] [1,4]benzodiazepines have a marked influence on the Ki(DI)/Ki(DS) ratios. In order to more precisely determine the structural requirements for both high affinity and selectivity at DI BzR relative to DS, 3D-QSAR analyses were carried out on ligand affinities at both of these BzR isoforms. This analysis was based, in part, on the new X-ray crystallographic data. Satisfactory cross-validated regression equations were obtained individually for the logarithms of ligand affinities at DI and DS as well as for the differences of the logarithms of their affinities at these two isoforms (cross-validated R2 > 0.70 for all three regression equations). The steric and electrostatic 3D-QSAR DI and DS maps are in qualitative accord with the structure-activity relationship (SAR) data. Furthermore, the DI and DI/DS maps may be useful in the design of ligands with enhanced DI affinity and DI/DS selectivity, respectively.

Animals↗
PubMed1993

Expression of Pseudomonas aeruginosa virulence genes requires cell-to-cell communication.

Pseudomonas aeruginosa is an opportunistic human pathogen that causes a variety of infections in immunocompromised hosts and individuals with cystic fibrosis. Expression of elastase, one of the virulence factors produced by this organism, requires the transcriptional activator LasR. Experiments with gene fusions show that gene lasl is essential for high expression of elastase. The lasl gene is involved in the synthesis of a diffusible molecule termed Pseudomonas autoinducer (PAI). PAI provides P. aeruginosa with a means of cell-to-cell communication that is required for the expression of virulence genes and may provide a target for therapeutic approaches.

Amino Acid Sequence↗
PubMed1993

A comparison of tumor and normal tissue microvascular hematocrits and red cell fluxes in a rat window chamber model.

This laboratory has previously used a window chamber model to measure red blood cell velocity in mammary tumors and normal granulation tissues of the F-344 rat. Because red cell flux and hematocrit more accurately reflect the oxygen carrying potential of blood, we used this model to measure these parameters. Red blood cells were labelled with fluorescein isothiocyanate, and 0.2 ml. packed cells were injected intravenously into rats bearing an 8 to 10 day old R-3230 mammary carcinoma. beta-phycoerythrin (0.15 mg.) was also injected and served as a plasma dye to outline the blood vessels. A sample of peripheral blood was then taken and analyzed by flow cytometry to determine the labeled fraction of red blood cells. Flowing tumor and normal tissue vessels were recorded onto a VCR, and these video images were used to determine vascular length and diameter, RBC flux and velocity, and hematocrit. Median vessel diameter and loge (red blood cell flux) were significantly greater in tumors than in normal tissues (p = 0.007 and p < 0.025, respectively). After controlling for these variables, the median tumor hematocrit of 19% was not significantly greater than the median normal tissue hematocrit of 15%. This technique provides a nontoxic and reproducible method that is now being used to assist in the in vivo definition of tumor oxygenation.

Animals↗
PubMed1993

Thrombomodulin activity and localization.

An overview on the properties, actions and localization of thrombomodulin (TM) in situations of tissue injury and in selected tumors is presented. The localization and activity of TM after injury to vascular endothelium shows that following balloon catheter denudation of the endothelium of the rabbit aorta, the activity and immunohistochemical staining is markedly reduced. The functional and antigenic levels approach the control levels approximately one week after the initial injury. The results suggest that the neointimal smooth muscle cells express TM. This phenotypic plasticity of the neointimal smooth muscle cells may be important in conferring thrombo-resistance to the lumenal lining cells of vessels after injury. Studies are also reviewed on the use of soluble recombinant TM to prevent thrombosis after ligature of vessels in an experimental model. Further characterization on the immunohistochemical distribution of TM in normal tissues and tumors shows that staining with a monoclonal anti TM antibody can be very useful in separating mesotheliomas from pulmonary adenocarcinomas. These studies may lead to insights concerning the role of TM in tissue-injury-repair and tissue differentiation.

Adenocarcinoma↗
PubMed1992

Predictive binding of beta-carboline inverse agonists and antagonists via the CoMFA/GOLPE approach.

The synthesis and affinities of six new 3-substituted beta-carbolines (6-10, 12) for the benzodiazepine receptor (BzR) are described. These analogs were used both to probe the dimensions of the hydrophobic pocket in the benzodiazepine receptor and to test the predictive ability of a previously reported 3D-QSAR regression model. Of the new analogs synthesized, the gamma-branched derivatives (isobutoxy, 7, IC50 = 93 nM; isopentoxy, 9, IC50 = 104 nM) display significantly higher affinity for the BzR than either the beta-branched (sec-butoxy, 6, IC50 = 471 nM; tert-butyl ketone, 12, IC50 = 358 nM) or delta-branched (isopentoxy, 8, IC50 = 535 nM) analogs. An exception to this rule is the gamma-branched 3-benzyloxy derivative 10 (IC50 > 1000 nM) which appears to have a chain length that is too long to be accommodated by the BzR. The standard error of prediction for these six new beta-carbolines using the original regression model is significantly lower than the standard error estimate of the cross validation runs on the training set, hence the predictions made using this model are much better than expected. In order to obtain more credible predictions, a new procedure called GOLPE (generating optimal linear PLS estimates) was used to eliminate irrelevant electrostatic and steric descriptors from the regression equation. A substantial reduction in the standard error estimate resulted. The predictions from this new regression equation were somewhat less accurate than the ones obtained with the original regression equation; however the standard error of prediction and the standard error estimate are in much closer agreement. Finally, to probe the effect that the quality of the steric and electrostatic potentials has on 3D-QSAR analyses, the semiempirical MNDO parallel PRDDOE geometries and Mulliken charges used in the original analyses were replaced with ab initio 3-21G parallel 6-31G* geometries and electrostatic potential fit charges. A modest decrease in the standard error estimate and increase in cross validated R2 resulted.

Animals↗
PubMed1992

Synthesis of novel 2-phenyl-2H-pyrazolo[4,3-c]isoquinolin-3-ols: topological comparisons with analogues of 2-phenyl-2,5-dihydropyrazolo[4,3-c]quinolin-3(3H)-ones at benzodiazepine receptors.

Based on the topology of pyrazoloquinolinones 10-12, a series of 2-phenyl-2H-pyrazolo[4,3-c]isoquinolines 6a-d, 7a-d, 8, and 9 have been synthesized and evaluated for their ability to inhibit radioligand binding to benzodiazepine receptors (BzR). Modification of the hydrogen bonding donor and acceptor characteristics of the NH and C = O functionalities of the pyrazoloquinolinones 10-12 resulted in ligands with dramatically reduced affinities (IC50 much greater than 2 microM) for BzR. The low affinities of 6a-d, 7a-d, 8, and 9 are consistent with the involvement of the NH function present on diverse classes of inverse agonists (beta-carbolines, diindoles, and pyrazoloquinolinones) with a hydrogen bond acceptor site (A2) on the binding protein. Moreover, it supports the involvement of the carbonyl function of the pyrazoloquinolinones and the pyridine nitrogen atom of beta-carbolines and diindoles with a hydrogen bond donor site (H1). Finally, the results from this work indicate that a simultaneous interaction at both hydrogen bond donor (H1) and acceptor sites (A2) at BzR is required for high affinity binding of inverse agonists.

Animals↗
PubMed1992

Structure-function analysis of exotoxin A proteins with mutations at histidine 426.

Substitution of Tyr for His-426 of Pseudomonas aeruginosa exotoxin A results in a mutant protein with reduced ADP-ribosyltransferase activity (M. J. Wick and B. H. Iglewski, J. Bacteriol. 170:5385-5388, 1988). To investigate the role of His-426 in enzymatic activity, oligonucleotide-directed mutagenesis was used to construct mutant proteins encoding Ala, Glu, Gly, Lys, or Pro at position 426. The effect of these amino acid substitutions on ADP-ribosyltransferase activity was analyzed in 34,000-Da carboxy-terminal exotoxin A peptides (H426n peptides). ADP-ribosyltransferase activity of the H426n peptides fell within a range between 0.002 and 28% of wild-type levels of activity, suggesting that His-426 is required for full expression of enzymatic activity of exotoxin A. To investigate a possible catalytic function of His-426, the abilities of full-size (66,000-Da) wild-type exotoxin A and mutant proteins encoding either Ala-426 or Tyr-426 to hydrolyze NAD were compared by measuring NAD-glycohydrolase activity. This analysis revealed that exotoxin A encoding either Ala-426 or Tyr-426 expressed less than 1% of wild-type levels of NAD-glycohydrolase activity. Several criteria, including differential enzymatic activation properties and unique tryptic digestion patterns, revealed that the wild-type and mutant full-size proteins exhibit conformational differences. Our data suggest that His-426 plays a critical structural role in establishing the molecular architecture of the catalytic site in domain III and is important in orienting active-site residues in the cleft.

ADP Ribose Transferases↗
PubMed1992

Punishment of schedule-controlled behavior with beta-carboline injections: antagonism and comparisons with other compounds.

Squirrel monkeys were trained to press a key under a multiple schedule of food presentation. In the presence of either green or red stimulus lights, the 30th response produced a food pellet (fixed-ratio schedule). In the presence of the red stimulus lights (punishment component), the first response of each fixed-ratio produced either an i.v. injection of histamine [30.0-100.0 micrograms/kg/injection (inj)] or saline, accompanied by a 200-msec presentation of amber stimulus lights. Sessions in which histamine was injected alternated with sessions in which saline was injected. Another group of subjects was studied under identical schedule conditions except that electric shock was scheduled with the 200-msec stimulus light. During alternate sessions, electric shock at a high or low intensity with the stimulus, or the stimulus alone was scheduled. When performances stabilized, histamine or high intensity electric shock selectively suppressed responding in the punishment component; saline, low intensity electric shock or the stimulus light alone had no effects. Subsequently, different doses of histamine, I-nicotine, cocaine or beta-carboline-3-carboxylic acid ethyl ester (beta-CCE) were substituted for histamine during single sessions. Histamine (17.8-100 micrograms/kg/inj), I-nicotine (32 micrograms/kg/inj) and beta-CCE (10-56 micrograms/kg/inj), but not cocaine (10.0-100.0 micrograms/kg/inj), produced a dose-related selective suppression of responding similar to that obtained with electric shock, suggesting that the drugs were functioning as punishers. Punishment by beta-CCE was antagonized with the benzodiazepine antagonist, flumazenil.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Behavioral effects of benzodiazepine ligands in non-dependent, diazepam-dependent and diazepam-withdrawn baboons.

Acute i.m. injections of benzodiazepine receptor ligands were administered to baboons before 1-h observational sessions. The agonist midazolam produced sedative effects, the antagonist flumazenil produced no behavioral effects, the inverse agonist FG7142 produced tremor and the inverse agonist 3-carboethoxy-beta-carboline hydrochloride (beta CCE) produced tremor, vomiting, jerks and seizures. Co-administration of these drugs (midazolam + beta CCE, midazolam + flumazenil or flumazenil + beta CCE) produced a mutual antagonism of these effects. Compared to the non-dependent condition, in the diazepam-dependent condition (baboons maintained on 20 mg/kg per day diazepam) and withdrawn condition (dependent baboons tested during withdrawal), midazolam produced decreased sedative effects, flumazenil produced increased effects (i.e., tremor, vomiting and jerks), and beta CCE produced increased frequency of seizures. Taken together, these data suggest that (1) benzodiazepine receptor ligands lie on a continuum of behavioral activity, and (2) chronic diazepam administration alters the behavioral effects of these benzodiazepine ligands, producing a shift in the direction of the inverse agonist.

Animals↗
PubMed1991

Hyperparathyroidism presenting as an unusual cause of lower extremity gangrene.

Vascular surgeons are often consulted on the management of lower extremity ulcerations and gangrene which are commonly due to atherosclerotic arterial occlusive disease or to chronic venous insufficiency. The following report describes an unusual case of severe lower extremity cutaneous gangrene associated with secondary hyperparathyroidism and a review of the literature.

Amputation, Surgical↗
PubMed1991

Synthetic and computer assisted analysis of the pharmacophore for agonists at benzodiazepine receptors.

In order to employ rational drug design in the discovery of selective benzodiazepine receptor agonists and inverse agonists, pharmacophore/receptor models for both these activities must first be established. Recently, a pharmacophore for the inverse agonist site has been formulated employing the most recent receptor mapping techniques (22). The continuation of this approach to the pharmacophore for agonist ligands has permitted a definition of this site independently of the inverse agonist model. The agonist pharmacophore/receptor contains two hydrogen bond donating sites of interaction (H1 and H2) located about 6.5 A from each other, as well as three areas of lipophilic interaction (L1-L3). The areas L1 and L2 are critical for agonist activity; moreover, some ligands also require an interaction in a third lipophilic area termed L3. This is in agreement with previous work (12-23). In addition, an area of negative steric interaction (S1) between the ligand and receptor-binding protein is defined. In regard to the pharmacophore, it was established that the alignment rule for agonist beta-carbolines is different from that which elicits inverse agonist activity. Consideration of the pharmacophore has resulted in the synthesis of a new beta-carboline 16 which elicits agonist activity. This ligand 16 not only satisfied the requirements of the pharmacophore, but more importantly it elicited both anticonvulsant and anxiolytic activity, but was devoid of the myorelaxant/ataxic properties associated with the benzodiazepines.

Animals↗
PubMed1991

Thrombomodulin activity on human saphenous vein grafts prepared for coronary artery bypass.

Thrombomodulin, a membrane glycoprotein present on normal vascular endothelium, binds circulating thrombin and is important in protein C activation. These functions contribute to the nonthrombogenic nature of endothelium. Damage during harvest and ex vivo storage of vein grafts may result in dysfunction of this endothelial anticoagulant barrier and possibly contribute to early graft thrombosis. We studied the functional activity and antigenic expression of thrombomodulin on saphenous veins before (initial) and after (harvested) harvest and storage for coronary artery bypass grafting in 15 patients. Also, fresh saphenous vein was studied after mechanical endothelial stripping. After storage for 2.7 +/- 0.6 hours at room temperature in heparinized saline, thrombomodulin functional activity in harvested vein segments was 28% less than initial segments (p = 0.08). Endothelial stripping resulted in a 79% reduction in thrombomodulin activity compared with initial segments (p = 0.04). Immunohistochemical staining confirmed thrombomodulin antigen on vein grafts after harvest and storage, but not on segments stripped of endothelium. Thrombomodulin functional activity and antigenic expression on human saphenous vein grafts is not significantly changed by harvest and relatively short periods of storage at room temperature in heparinized saline.

Antigens↗